Collagen-like peptide bioconjugates - Bioconjugate Chemistry (ACS

Subscriber access provided by Fudan University

Review

Collagen-like peptide bioconjugates Tianzhi Luo, and Kristi L. Kiick Bioconjugate Chem., Just Accepted Manuscript • DOI: 10.1021/acs.bioconjchem.6b00673 • Publication Date (Web): 26 Jan 2017 Downloaded from http://pubs.acs.org on January 31, 2017

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

Bioconjugate Chemistry is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Bioconjugate Chemistry

Collagen-like peptide bioconjugates Tianzhi Luoa, Kristi L. Kiicka,b,c,* a

Department of Materials Science and Engineering, University of Delaware, Newark, DE, 19716, USA b

Department of Biomedical Engineering, University of Delaware, Newark, DE, 19716, USA c

Delaware Biotechnology Institute, Newark, DE, 19711, USA.

Abstract Collagen-like peptides (CLPs), also known as collagen-mimetic peptides (CMPs), are short synthetic peptides that mimic the triple helical conformation of native collagens. Traditionally, CLPs have been widely used in deciphering the chemical basis for collagen triple helix stabilization, mimicking collagen fibril formation and fabricating other higher-order supramolecular self-assembles. While CLPs have been used extensively for elucidation of the assembly of native collagens, less work has been reported on the use of CLP-polymer and CLP-peptide conjugates in the production of responsive assemblies. CLP triple helices have been used as physical cross-links in CLP-polymer hydrogels with predesigned thermoresponsiveness. The more recently reported ability of CLP to target native collagens via triple helix hybridization has further inspired the production of CLPpolymer and CLP-peptide bioconjugates and the employment of these conjugates in generating well defined nanostructures for targeting collagen substrates. This review summarizes the current progress and potential of using CLPs in biomedical arenas and is intended to serve as a general guide for designing CLP-containing biomaterials.

1. Introduction Collagen is the main component of the extracellular matrix (ECM) and comprises 25% to 35% of protein in humans. Collagens are found mainly as fibrous proteins (type I, II and III) located in tendon, ligament, skin, cartilage and bone. Other types of collagens, such as type IV and type VIII collagens, are essential in the formation of network structures such as basement membranes.1 With a total of at least 28 different types discovered, collagens play vital roles in biological activities such as mediating cell adhesion, cell migration, tissue scaffolding and tissue repair.1 Although the architectures and roles of these collagens vary widely, they all share the same tertiary structure – the collagen triple helix.2, 3 The collagen triple helix comprises three polyproline-II-type helices, twisted together in a right-handed form. Endowed with outstanding gel-forming abilities and biodegradability, collagens have been widely utilized for drug delivery, cell culture, tissue support and regeneration.1, 4-9 However, some of the restrictions of using animal-derived collagens, including thermal

ACS Paragon Plus Environment


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

instability and possible contamination with pathogenic substances10 have inspired the development of short synthetic model collagens, widely known as collagen-like peptides (CLPs) or collagen-mimetic peptides (CMPs). CLPs are short synthetic peptides that possess the triple helical conformation of native collagens. The canonical collagen repetitive motif comprises glycine (Gly) - X - Y tripeptide repeats, where X and Y residues are generally proline (Pro) and hydroxyproline (Hyp). In the past few decades, collagens and collagen-like peptides have been widely demonstrated to have unique structure and properties. Traditionally, CLPs have been widely used as model peptides to study the triple helix stability of native collagens11 and to direct self-assembly into higher-order structures.12, 13 The use of collagen-like peptides as bioactive domains and physical cross-linkers in hydrogels14, 15 have illustrated their potential to improve cell adhesion, proliferation and ECM production. More recently, CLPs have also been conjugated to synthetic polymers and other (poly)peptides, to fabricate thermoresponsive bioconjugates for tissue engineering and targeted drug delivery purposes.16-18 This review summarizes current progress and potential of using CLPs in these fields. Readers interested in biological application of native collagens are directed to previously published reviews.6-9, 19 2. Triple helix stability In order to utilize CLPs in fabricating biomaterials or as labelling agents, drug delivery vehicles and tissue engineering scaffolds, it has been essential to understand the sequence-based thermal stability of the triple helix and other factors affecting it. In the early stages of such work, Brodsky and coworkers investigated the influence of guest residues at X and Y positions using Gly-X-Hyp and Gly-Pro-Y as guest sequences in the context of the (Gly-Pro-Hyp)8 host peptide and reported the strength of each of the natural amino acids in stabilizing the collagen triple helix.11 Since then, a wide variety of methods aimed at improving the stability of CLP triple helices have been deployed, which includes but is not limited to: i) replacing hydroxyproline with amino acids with high electronegative Cγ substitutions such as fluoride and chloride;20-27 ii) introducing one-residue staggered pairwise electrostatic interactions,28, 29 either within the same GXY triplets,28 or between adjacent triplets;30-32 iii) functionalizing the terminus of CLP with ligands for metal ion coordination;33, 34 iv) introducing a collagen type III derived cysteine knot at the C-terminus;35-39 v) linking three α-chains covalently at the C-terminus using templates such as cis,cis-1,3,541 trimethylcyclohexane-1,3,5-tricarboxylic acid (Kemp triacid),40, tris(241 aminoethyl)amine-(suc-OH)3 (TREN-(suc-OH)3) and the N-terminus of the 42 bacteriophage T4 fibritin foldon domain; and vi) introducing strong hydrophobic interactions using alkyl chain tails.43-45 The details of these designs have been presented in a previous review.46 More recently, our group has designed CLP-conjugates with thermoresponsive polymer poly(diethylene glycol methyl ether methacrylate) (PDEGMEMA), as well as thermoresponsive elastin-like peptides (ELPs).16-18 Circular


Page 2 of 23

Page 3 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


dichroism spectroscopy results show that the collapsed polymer/peptide domain also can act as an anchoring point to stabilize the triple helix. Despite the various methods aimed at modifying the side chain and terminus of the CLP, the backbone of the peptide, especially the glycine residue, has remained largely intolerant to any substitution. By substituting one amide bond of the peptide backbone using a thioamide, Raines and coworkers reported a backbone-modified collagen triple helix that does not suffer a loss in thermostability.47 More recently, by simply replacing the α-carbon of one glycine in (GPO)7 with a nitrogen atom, Chenoweth and coworkers observed a significant increase in Tm of ∼10 °C compared to the unsubstituted CLP.48 Based on molecular dynamics calculations, the hyper stability of the triple helix was attributed to the formation of additional hydrogen bonds from azaglycine (azGly) to carbonyls on adjacent peptide strands in the helix. Continued work from the same group49 suggested that the substituted azGly had a positional preference, with a slightly higher Tm for CLPs with a central substitution over N- or C-terminal substitutions. Additionally, the inclusion of multiple azGly residues in the CLP led to a synergistic stabilization effect. As a result, a CLP with a sequence as short as (azGly-Pro-Hyp)4 was able to selfassemble into a defined protein tertiary structure at physiological temperature as assessed via circular dichroism spectroscopy, although further investigation is necessary to confirm if this structure is strictly collagen-like triple helix or some other trimeric structure. These new approaches suggest that nature’s building blocks may not optimize the stability of collagen triple helix, and by introducing precisely designed artificial amino acids into the backbone, triple helix stability may be increased. Additionally, the introduction of these artificial amino acids also decreases the proteolytic degradation of the modified peptides/proteins,50, 51 which may be utilized to improve the stability of such biomaterials for in vivo studies. 3. Higher-order assembly of CLPs Collagen-like peptides as simple as (Pro-Hyp-Gly)10 are observed to form higher order structures52 via a nucleation and growth mechanism. Synthetic fibers self-assembled from CLPs, which mimic native collagen fibers, have been produced successfully via end-toend interactions of various well-designed CLP systems. For example, Maryanoff and coworkers reported micrometer-scale fibrillar structures assembled through noncovalent CLP end-to-end π-π interactions.12, 13 By introducing lysine residues into this CLP sequence, Raines and coworkers53 decorated collagen-like fibers with gold nanoparticles that were fashioned into electrically conductive metal nanowires by metallization through electroless silver plating (Figure 1a). In addition to π-π stacking, some other interactions have also been reported for the construction of collagen-like fibrillar structures, such as self-complementary ligation between CLPs functionalized with a staggered cysteine knot,54-56 cation-π interactions between positively charged N-terminal Arg residues and C-terminal Phe residues,57 CH-π interactions between imino acids (Pro and Hyp) and aromatic residues (Phe and Tyr),58 as well as lateral electrostatic attractions between



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

positively and negatively charged CLP domains (Figure 1b).59-61 Readers interested in details of self-assembly using these strategies are directed to previous reviews.46, 62 More recently, Zhang and coworkers functionalized the hydroxyl groups of hydroxyproline of the CLP with linear or dendritic oligoethylene glycols (OEG).63 The resulting CLPs were observed to form long fibers, possibly driven by the radial amphiphilicity of the solvophilic dendrons on the periphery and solvophobic polypeptide backbone in the center (Figure 1c).

Figure 1. Mimicking collagen fibers via CLP self-assembly. a) left: Collagen-like fibers used as templates for the assembly of metallic nanowires; right: TEM image of goldlabeled CLP fibers after electroless silver plating; b) Self-assembly of collagen fibers with well-defined D-periodicity using axially staggered interchain electrostatic interactions; c) Molecular structure and fibril formation of oligoethylene glycol (OEG)functionalized CLP. (Reproduced with permission from Ref 53, 59, 63.)


Page 4 of 23

Page 5 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


In addition to interactions between the CLP chains, coordination bonds between metal ions and specific ligands have also been introduced for controlled self-assembly of CLPs. Our previously published review46 discussed a coassembly system designed by Chmielewski and coworkers, which involved metal ions and CLPs functionalized with metal-binding units at both termini.64-66 Such metal binding ligands have also been conjugated to a centrally positioned sidechain of the CLP.67 TEM results suggested that subsequent addition of metal ions triggered radial growth of the triple helices into fibrous structures (Figure 2a). Although the assembled fibers lack the periodic bands of native collagen, these data provide proof that collagen-like fibers can be generated via nonlinear assembly. More recently, researchers from the same group extended this work by introducing three ligands to the CLP.68 The resulting triple helix, which radially displayed nine hydrophobic bipyridine moieties, was reported to self-assemble into micrometer-scaled disks with a curved morphology (Figure 2b). More interestingly, a morphological transition from curved disks to micrometer-sized hollow spheres was observed after the addition of metal ions. Owing to the two-step formation of these microspheres, fluorescently labeled dextrans were able to be encapsulated into these CLP peptide microcages under mild conditions (Figure 2c).69 Temperature-controlled release of the cargo was then achieved by unfolding the peptide cage above the Tm of the CLP. The reported hierarchical assembly of these ligand-equipped CLP into triple helices, disks, and metal-promoted microcages was reminiscent of the assembly of a number of viral, bacterial and protein cages,69 although the sizes of the CLP-derived cages were about one to two orders of magnitude larger than those observed in nature. The mild loading conditions and thermoresponsive release of encapsulated dextran suggests the potential for these peptide cages in the delivery of biopolymers.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Figure 2. Metal-assisted self-assembly of CLPs. a) Radial assembly of collagen peptide fibers from metal-ligand coordination; b) Ligand functionalized CLP triple helices selfassemble into curved disks, followed by metal-promoted assembly into hollow spheres; c) Encapsulation of fluorescein-labeled dextran within the interior of CLP assembled cages (left), with the exteriors stained with Congo Red (right) and the overlap of the green and red channels (center) (scale bar = 1 µm). (Reproduced with permission from Ref 67-69.) 4. Targeting native collagen via triple helix hybridization In addition to the majority of CLP research on triple helix stability and higher-order assembly, Yu and coworkers have made important contributions to utilizing the binding interactions between CLP and native collagens for a range of applications, based on their observation that unfolded single-strand collagen-like peptides have a strong propensity to bind to native collagen via a strand hybridization process (Figure 3a).70 Based on this physical hybridization, fluorescently labeled CLPs have been used as collagen-specific stains in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) for direct detection of various types of collagens including intact type I, II, IV collagen, as well as MMP-cleaved type I collagen.71 Collagen bands containing as little as 5 ng of protein were successfully detected, suggesting a high staining efficiency. These


Page 6 of 23

Page 7 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


fluorescently labeled CLPs have also been utilized as staining reagents for native collagens in human tissues including skin, cornea, bone71 and liver.72 Images of CLPstained human liver tissue were almost identical to images stained by collagen antibodies (Figure 3b).72 The binding interaction between CLP and native collagen has also been used for encoding cellular cues in collagen scaffolds. For example, studies from the same group73 reported a new collagen-like peptide consisting of eight negatively charged glutamic acid residues at the N-terminus of (Pro-Hyp-Gly)9. The anionic CLP was not only able to bind to type I collagen via strand-strand hybridization, but also able to attract vascular endothelial growth factors (VEGFs) through electrostatic interactions, which then induced tubulogenesis of endothelial cells (ECs) pre-encapsulated within the collagen substrate. In order to test the ability of the CLP to target pathological tissues of high MMP activity, in vivo tumor-targeting experiments were conducted on mice bearing subcutaneous PC-3 prostate tumor xenografts.74 The fluorescently labeled CLP was able to permeate the tumor vasculature and accumulate at the tumor sites (Figure 3c). A high level of accumulation of the CLP was also observed within the skeleton and joints, especially in regions with high MMP and collagen remodeling activity such as articular cartilage of the knee (Figure 3d). As the peptide can be integrated, with high stability, into native collagen both in vitro and in vivo, these research activities suggest that approaches which employ CLPs should thus offer particular promise for biomedical applications such as tumor, bone and cartilage imaging, as well as targeted drug delivery into these regions. The triple helix hybridization has also been employed to impart collagen-targeting properties to preformed nanoparticles.75-77 For example, gold nanoparticles (Au-NP) conjugated with short CLPs showed specific binding with reconstituted type I collagen fibers (Figure 3e), while Au-NP functionalized with a scrambled G7P7O7 peptide, incapable for forming triple helix, exhibited little binding with the collagen fibers.75 Two types of Au-NPs with different diameters, which were decorated with parallel CLP strands (via either the N- or C-termini), formed structured aggregates, presumably attributed to inter-particle triple helix formation (Figure 3f).76 Additionally, larger CLP Au-NPs with diameters of approximately 100 nm were shown to be competent for extracting gelatin from protein mixtures. Considering the wide utilization of gelatin as a stabilizer additive for protein reagents and therapeutics, these CLP-NPs could be used for removal of gelatin from such formulations to minimize side effects.76



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Figure 3. Targeting native collagens via triple helix hybridization. a) Schematic of CLP strand hybridization process; b) CLP labels collagens of human liver tissue as efficiently as collagen antibodies; c) In vivo NIRF images of mice bearing PC-3 prostate tumors administered with fluorescent labelled (GPO)9 CLP indicate specific tumor accumulation; d) Dual-NIRF image of the knee joint showing the uptake of fluorescent labelled CLP (red) and BoneTag™ (stains calcifying tissues in green), CLP-specific uptake can be seen within the articular cartilage and meniscus (red arrowhead) as well as focal regions within the tibia and the femur head; e) TEM micrographs of reconstituted type I collagen fibers labelled with CLP-functionalized gold nanoparticles; f) left: schematic of assembly from gold nanoparticles functionalized with parallel CLP strands of which N- or Cterminal was conjugated; right: TEM image of the assembled CLP-NPs. (Reproduced with permission from Ref 72, 74-76.) Inspired by these studies, the Kiick and Sullivan laboratories have employed CLPs to control the retention and delivery of DNA polyplexes from collagen structures, including both 2-D collagen films and 3-D hydrogels (Figure 4a).78 The retention and release of the DNA polyplex on collagen substrates is easily tuned by varying the concentration of CLP displayed on the polyplex. The results indicate substantially improved retention of the CLP-modified polyplexes compared with unmodified polyplexes (Figure 4b). The CLPmodification of DNA polyplex has also been shown to enhance the expression of encoded platelet-derived growth factor–BB (PDGF-BB) genes, supporting enhanced wound closure rates in an in vitro cell-based wound-closure model (Figure 4c).79 These


Page 8 of 23

Page 9 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


results suggest that this technique may be used to create tunable, collagen-based delivery systems for wound repair and other regenerative medicine applications.

Figure 4. CLP-modified DNA polyplexes for modifying collagen scaffolds. a) Schematic illustration of targeting type I collagen scaffold using CLP-modified DNA-polyplex; b) Initial retention of DNA on collagen films as a function of the percent CLP-PEI in the polyplex; c) In vitro wound model. Left: defects in cell-seeded collagen gels were filled with collagen scaffolds modified with rPDGF-BB, polyplex encoding for luciferase, or polyplex encoding for PDGF-BB, and subsequent defect invasion or “wound closure” was monitored via microscopy. Scale bars = 250 µm; right: quantitative analysis of the wound closure. (Reproduced with permission from Ref 78, 79.)

5. CLP-containing hydrogels Collagen-like peptides have also been used to fabricate scaffolds for tissue engineering applications. In one of the earliest approaches to such applications, Hartgerink and coworkers reported CLP hydrogels stabilized by triple helical lateral electrostatic interactions (Figure 5a).60 Rheology results indicated the high storage modulus (G’) of the hydrogel, comparable to native collagen hydrogels, as well as the degradation of the hydrogels by collagenase at a similar rate to the degradation of native rat-tail collagen. Given their triple helix formation, CLPs can also be utilized as physical cross-linkers to generate hydrogels with well controlled thermoresponsiveness. For example, multi-armed PEG hydrogels functionalized with CLP chains (Figure 5b) have been reported by the Yu



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

research group,15 as well as the Chmielewski research group.80 As expected, the thermal stability of the gels can be easily controlled by changing the length of the CLP. More interestingly, the mechanical properties of these gels could also be manipulated via the addition of free CLPs, which competed as binding partners with the CLPs in the crosslinks. More recently, the biocompatibility of these PEG-CLP hydrogels was tested by Phopase and coworkers.81 In vitro culturing of human corneal epithelial cells in the hydrogel showed good proliferation without any cytotoxic effects (Figure 5c, left). After 90 days of implantation into the dorsum of rats, the hydrogel was relatively intact, and remained free of immune cells (Figure 5c, right), indicating the compatibility of the PEGCLP in vivo. More importantly, when tested preclinically as corneal implants in mini-pigs, the PEG-CLP implants remained stably incorporated and optically clear after 12 months of implantation, similar to the contralateral unoperated control corneas (Figure 5d), suggesting the potential utility of these PEG-CLP based hydrogels as alternatives to human donor corneal transplantation. In addition to physical cross-linkers, CLPs integrated with certain bioactive domains have also been widely used to fabricate hydrogels to improve cell adhesion and ECM production.82, 83 For example, Yu and coworkers recently reported a CLP-conjugated poly(ethylene glycol) diacrylate (PEODA) hydrogel encoded with RGD cell binding domain,14 which showed improved adhesion and proliferation of encapsulated fibroblasts. Tong and coworkers84 synthesized a CLP amphiphile with a C16 lipophilic tail attached to the C-terminus of a CLP with sequence (GPO)3GFOGER(GPO)3. Given the inclusion of the integrin-specific binding sequence GFOGER, hydrogels formed from the amphiphile were shown to promote the adhesion and spreading of HepG2 cells. By introducing a hydroxyproline-free CLP comprising an integrin-binding GER triplet, our group successfully promoted integrin-mediated adhesion, spreading and proliferation of human mesenchymal stem cells (hMSCs) in a hyaluronic acid (HA) particle-based hydrogel system.85 The integration of integrin binding domains with these CLPcontaining hydrogels shows great potential for the production of biocompatible substrates for cell proliferation and tissue regeneration applications. Building upon the utility of metal-ligand coordination interactions mentioned above, Chmielewski and coworkers designed CLP sequences equipped with bipyridine metal ligands and RGDS cell adhesion domain.86 The CLPs assembled into 3-D scaffolds upon the addition of metal ions, with the encapsulation of coassembled His-tagged cargoes including green fluorescent protein (GFP-His8) and human epidermal growth factor (hEGF-His6) (Figure 5e, left). The bound hEGF-His6 was found to be gradually released from the matrix in vitro and to induce both cell proliferation and the organization of a human non-tumorigenic epithelial cell line (MCF10A) into spheroid structures (Figure 5e, right). The ability to capture in these materials some of the early events of MCF10A acini formation is likely to be useful for modeling morphogenesis and carcinogenesis in vitro.


Page 10 of 23

Page 11 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 5. CLP-containing hydrogels. a) Self-assembly of the CLP (Pro-Lys-Gly)4-(ProHyp-Gly)4-(Asp-Hyp-Gly)4 into nanofibers and subsequent fabrication of CLP hydrogel at higher concentrations; scale bar = 1 µm; b) 4-arm PEG hydrogel crosslinked via (GPO)9 collagen triple helix interactions. Left bottom panel: storage and loss modulus for the hydrogel (10 wt%) before and after the addition of free CLP solution; c) Left: human corneal epithelial cells after 5 days in culture on PEG-CLP hydrogels, stained with live/dead stain; Right: representative H&E stained section of PEG-CLP hydrogel implanted subcutaneously into a rat for 90 days. Inset: intact 10 mm diameter, 500 µm thick sample retrieved after implantation. Scale bar = 50 µm; d) Left: cornea-shaped implant made from PEG-CLP hydrogel; middle: optically clear PEG-CLP hydrogel implant (arrowed) integrated within the pig cornea; right: healthy, unoperated contralateral cornea; e) Metal-promoted gelation of CLPs with the encapsulation of Histagged biomolecules. The hydrogel induced organization of the MCF10A cell line into spheroids. (Reproduced with permission from Ref 15, 60, 81, 86.) 6. CLP-polymer conjugates While peptide-based biomaterials exhibit well defined hierarchical structures and a variety of biological functions such as cell recognition, adhesion and proliferation,85, 87, 88 certain well controlled synthetic polymers have been observed to be highly biocompatible



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

and resistant to enzymatic degradation. Chemical conjugation of polypeptides or proteins with synthetic polymers thus has been widely used to combine the advantages of the two, via approaches including solid-phase peptide-coupling chemistry,89-91 Schiff base formation,92-95 Staudinger ligation,96 copper-catalyzed azide-alkyne cycloaddition (CuAAC) click reaction97-100 and Michael-type addition.101-104 The introduction of the efficient coupling chemistries discovered in the past few decades has given rise to an expansion in the design of novel nanometer- and micron-sized structures assembled from polymer-peptide bioconjugates, with promising chemical and biological functions.105, 106 As comprehensively reviewed, a variety of peptides including α-helical peptide domains,107-111 β-sheet peptide motifs,112-115 and elastin mimetic peptides116-122 have been successfully employed to produce these hybrid materials. Compared with the surge in research on these peptide–polymer conjugates, collagen-like peptide hybrid materials represent a relatively new area. Tirrell and coworkers44 attached C12 head group to a collagen-like peptide and found that the conjugation with a hydrophobic C-terminal head group improved the thermal stability of the collagen triple helix. Tong and coworkers84 reported that after a C16 lipophilic tail was attached to the C-terminus of a CLP with the sequence (GPO)3GFOGER(GPO)3, the amphiphile selfassembled into micrometer-long nanofibers with a diameter of approximately 16 nm under aqueous conditions (Figure 6a). These results demonstrated that after conjugation with a more hydrophobic moiety, the triple helix forming CLPs can still be used as building block for the assembly of higher-order structures. More recently, our group, in collaboration with Theato and coworkers, has conjugated CLPs with the thermoresponsive polymer poly (diethylene glycol methyl ether methacrylate) (PDEGMEMA), and studied the conformational and assembly behavior of the resulting PDEGMEMA-CLP diblock17 or PDEGMEMA-CLP-PDEGMEMA triblock.18 For the triblock, the CLP domain adopts a triple helix conformation after conjugation with the polymer, with electron microscopy indicating that the collapse of polymeric domain at 37 °C causes the formation of large hollow spherical structures (Figure 6b) that undergo an additional morphological transformation into fibrils with the elevation of temperature to 75 °C (Figure 6b), likely due to the unfolding of the collagen triple helix domain. DLS results suggested the PDEGMEMA-CLP diblock similarly exhibited a transition temperature around 32 °C (Figure 6c), above which the polymer domain collapsed and triggered the reversible diblock self-assembly into well-defined vesicles with diameters of approximately 50–200 nm (Figure 6d). The thickness of the vesicle surfaces was approximately 10–20 nm, consistent with the calculated thickness of a diblock bilayer. Due to the strong propensity for CLPs to bind to native collagen via strand hybridization processes discussed in the previous section, these nanosized vesicles may have significant potential as drug carriers for targeted delivery.


Page 12 of 23

Page 13 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Figure 6. Nanostructures from CLP-polymer conjugates a) TEM image of self-assembled nanofibers from CLP with sequence (GPO)3GFOGER(GPO)3 functionalized with C16 lipophilic tail; b) Left: Cryo-SEM image of spherical structures self-assembled from PDEGMEMA-CLP-PDEGMEMA triblock at 37 °C. Scale bar = 50 µm; right: TEM image of collagen like fibrils at 75 °C. Scale bar = 0.5 µm; c) Study of transition temperature of PDEGMEMA-CLP diblock via dynamic light scattering suggested an LCST of approximately 32 °C; d) TEM images of nanovesicles formed from PDEGMEMA–CLP diblock. (Reproduced with permission from Ref 17, 18, 84.) 7. CLP-ELP bioconjugates Elastin is an important component of extracellular matrix (ECM), especially in connective tissues. Tropoelastin, the soluble precursor of elastin, comprises alternating hydrophobic and hydrophilic domains, and elastin-like polypeptides (ELPs) composed of the hydrophobic pentapeptide Val-Pro-Gly-Xaa-Gly have been widely reported.123 The characteristic, LCST-like inverse temperature transition of this class of polypeptides119, 124 endows ELPs with the ability to be used as building blocks for thermoresponsive smart biomaterials.125, 126 In addition to the ELP block copolypeptides studied by Chilkoti,127-131 Conticello,132-135 Montclare,136-140 and others,141 ELPs have also been widely conjugated to synthetic polymers to generate thermoresponsive assemblies.142-144 Although ELPs have been widely utilized as bioconjugates with desired thermoresponsiveness in the past few decades, essentially all the ELPs employed have been recombinant, comprising tens or even hundreds of pentapeptide repeats. Short synthetic ELPs (e.g., those with fewer than 10 pentapeptides) have not been used for the fabrication of thermoresponsive nanoparticles, likely due to their extremely high



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

transition temperatures.145, 146 Most recently, our group has successfully overcome this limitation by conjugating short ELPs with a collagen-like peptide.16 According to DLS results, after conjugation with CLP, the inverse transition temperature (Tt) of ELP-CLP diblock conjugate dropped more than 80 °C, leading to the self-assembly of nanovesicles at temperatures as low as 4 °C (Figure 7). This unexpected reduction in LCST is almost certainly attributable to the anchoring effects of the CLP triple helix, which serves to locally isolate three ELP domains at concentrations approximately 100-fold higher than that of the ELP monomers in solution. More interestingly, further heating of the nanoparticles, to temperatures above the melting temperature of the CLP triple helix, fully resolubilized the nanoparticles into monomers (Figure 7), corroborating the hypothesis that the reduction of Tt was derived from the triple helix anchoring effect. Given the above-mentioned ability of CLPs to target native collagens via triple helix hybridization, our results not only provide a simple and versatile avenue to employ short synthetic ELPs for the fabrication of drug delivery vehicles, but also suggest future opportunities for targeting these drug carriers to collagen-containing substrates, relevant for a wide range of collagen-associated diseases.74, 147-153

Figure 7. Noncovalent modulation of the inverse temperature transition and self-assembly of ELP-CLP bioconjugates. a) Hydrodynamic diameter of nanostructures as a function of temperature upon heating; b) left: TEM image of nanoparticles formed at 37 °C, with an inserted cryo-TEM micrograph showing the vesicular structure; right: nanoparticles are resolubilized at 80 °C; c) proposed assembly mechanism of bilayer vesicles above the inverse transition temperature of ELP domain and subsequent disassembly of the nanoparticles above the melting temperature of CLP triple helix. (Reproduced with permission from Ref 16.) 8. Conclusions


Page 14 of 23

Page 15 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Collagen-like peptides and their conjugated hybrids have been widely demonstrated to have unique structure and properties. Traditional application of CLPs to decipher the role of natural amino acid residues and tripeptide motifs in stabilizing the collagen triple helix and mimicking collagen fibril formation has yielded much information about the assembly and activities of collagen. The expansion of these approaches by the introduction of specific interactions including electrostatic interactions, π-π stacking interactions and metal-ligand coordination, has introduced a variety of artificial collagenlike peptides with well-defined sequences that also form higher order assemblies. The use of collagen-like peptides as bioactive domains and physical cross-linkers in hydrogels, have illustrated their potential to improve cell adhesion, proliferation and ECM production. With the introduction of highly efficient coupling chemistries discovered in the past few decades, collagen-like peptides have also been conjugated with synthetic polymers and (poly)peptides, to generate stimuli responsive hybrid materials that are capable of self-assembling into well-defined nanostructures at physiological temperature. Although the application of these hybrids in specific biological applications remains in its early stages, the critical role of collagens in the extracellular matrix and the unique ability of CLP to target native collagens, suggests the importance of developing collagen-like containing biomaterials for targeted drug delivery and tissue engineering applications. References (1) (2) (3) (4)

(5)

(6) (7)

(8) (9) (10) (11) (12)

Gelse, K., Poschl, E., and Aigner, T. (2003) Collagens - structure, function, and biosynthesis. Adv. Drug Deliver. Rev. 55, 1531-1546. Ramachandran, G. N., and Kartha, G. (1955) Structure of collagen. Nature 176, 593-595. Okuyama, K. (2008) Revisiting the molecular structure of collagen. Connect. Tissue Res. 49, 299-310. von Arx, T., and Buser, D. (2006) Horizontal ridge augmentation using autogenous block grafts and the guided bone regeneration technique with collagen membranes: a clinical study with 42 patients. Clin. Oral Implant. Res. 17, 359-366. Haslik, W., Kamolz, L. P., Nathschlager, G., Andel, H., Meissl, G., and Frey, M. (2007) First experiences with the collagen-elastin matrix Matriderm((R)) as a dermal substitute in severe burn injuries of the hand. Burns 33, 364-368. Chattopadhyay, S., and Raines, R. T. (2014) Collagen-based biomaterials for wound healing. Biopolymers 101, 821-833. Abou Neel, E. A., Bozec, L., Knowles, J. C., Syed, O., Mudera, V., Day, R., and Hyun, J. K. (2013) Collagen - emerging collagen based therapies hit the patient. Adv. Drug Deliver. Rev. 65, 429-456. Ferreira, A. M., Gentile, P., Chiono, V., and Ciardelli, G. (2012) Collagen for bone tissue regeneration. Acta Biomater. 8, 3191-3200. Parenteau-Bareil, R., Gauvin, R., and Berthod, F. (2010) Collagen-based biomaterials for tissue engineering applications. Materials 3, 1863-1887. Nemoto, T., Horiuchi, M., Ishiguro, N., and Shinagawa, M. (1999) Detection methods of possible prion contaminants in collagen and gelatin. Arch. Virol. 144, 177-184. Persikov, A. V., Ramshaw, J. A. M., Kirkpatrick, A., and Brodsky, B. (2000) Amino acid propensities for the collagen triple-helix. Biochemistry 39, 14960-14967. Cejas, M. A., Kinnney, W. A., Chen, C., Vinter, J. G., Almond, H. R., Balss, K. M., Maryanoff, C. A., Schmidt, U., Breslav, M., Mahan, A., Lacy, E., and Maryanoff, B. E.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(13)

(14) (15)

(16)

(17)

(18)

(19) (20) (21) (22)

(23)

(24)

(25)

(26) (27)

(28)

(29)

(2008) Thrombogenic collagen-mimetic peptides: Self-assembly of triple helix-based fibrils driven by hydrophobic interactions. P. Natl. Acad. Sci. U.S.A. 105, 8513-8518. Cejas, M. A., Kinney, W. A., Chen, C., Leo, G. C., Tounge, B. A., Vinter, J. G., Joshi, P. P., and Maryanoff, B. E. (2007) Collagen-related peptides: Self-assembly of short, single strands into a functional biomaterial of micrometer scale. J. Am. Chem. Soc. 129, 2202-+. Stahl, P. J., and Yu, S. M. (2012) Encoding cell-instructive cues to PEG-based hydrogels via triple helical peptide assembly. Soft Matter 8, 10409-10418. Stahl, P. J., Romano, N. H., Wirtz, D., and Yu, S. M. (2010) PEG-based hydrogels with collagen mimetic peptide-mediated and tunable physical cross-links. Biomacromolecules 11, 2336-2344. Luo, T. Z., and Kiick, K. L. (2015) Noncovalent modulation of the inverse temperature transition and self-assembly of elastin-b-collagen-like peptide bioconjugates. J. Am. Chem. Soc. 137, 15362-15365. Luo, T. Z., He, L. R., Theato, P., and Kiick, K. L. (2015) Thermoresponsive selfassembly of nanostructures from a collagen-like peptide-containing diblock copolymer. Macromol. Biosci. 15, 111-123. Krishna, O. D., Wiss, K. T., Luo, T. Z., Pochan, D. J., Theato, P., and Kiick, K. L. (2012) Morphological transformations in a dually thermoresponsive coil-rod-coil bioconjugate. Soft Matter 8, 3832-3840. Lee, C. H., Singla, A., and Lee, Y. (2001) Biomedical applications of collagen. Int. J. Pharm. 221, 1-22. Holmgren, S. K., Taylor, K. M., Bretscher, L. E., and Raines, R. T. (1998) Code for collagen's stability deciphered. Nature 392, 666-667. Holmgren, S. K., Bretscher, L. E., Taylor, K. M., and Raines, R. T. (1999) A hyperstable collagen mimic. Chem. Biol. 6, 63-70. Eberhardt, E. S., Panasik, N., and Raines, R. T. (1996) Inductive effects on the energetics of prolyl peptide bond isomerization: Implications for collagen folding and stability. J. Am. Chem. Soc. 118, 12261-12266. Inouye, K., Sakakibara, S., and Prockop, D. J. (1976) Effects of stereo-configuration of hydroxyl group in 4-hydroxyproline on triple-helical structures formed by homogeneous peptides resembling collagen. Biochim. Biophys. Acta 420, 133-141. Bretscher, L. E., Jenkins, C. L., Taylor, K. M., DeRider, M. L., and Raines, R. T. (2001) Conformational stability of collagen relies on a stereoelectronic effect. J. Am. Chem. Soc. 123, 777-778. DeRider, M. L., Wilkens, S. J., Waddell, M. J., Bretscher, L. E., Weinhold, F., Raines, R. T., and Markley, J. L. (2002) Collagen stability: Insights from NMR spectroscopic and hybrid density functional computational investigations of the effect of electronegative substituents on prolyl ring conformations. J. Am. Chem. Soc. 124, 2497-2505. Shoulders, M. D., Guzei, I. A., and Raines, R. T. (2008) 4-chloroprolines: Synthesis, conformational analysis, and effect on the collagen triple helix. Biopolymers 89, 443-454. Kusebauch, U., Cadamuro, S. A., Musiol, H. J., Lenz, M. O., Wachtveitl, J., Moroder, L., and Renner, C. (2006) Photocontrolled folding and unfolding of a collagen triple helix. Angew. Chem. Int. Edit. 45, 7015-7018. Persikov, A. V., Ramshaw, J. A. M., Kirkpatrick, A., and Brodsky, B. (2002) Peptide investigations of pairwise interactions in the collagen triple-helix. J. Mol. Biol. 316, 385394. Kramer, R. Z., Venugopal, M. G., Bella, J., Mayville, P., Brodsky, B., and Berman, H. M. (2000) Staggered molecular packing in crystals of a collagen-like peptide with a single charged pair. J. Mol. Biol. 301, 1191-1205.


Page 16 of 23

Page 17 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(30)

(31)

(32)

(33) (34)

(35)

(36)

(37)

(38)

(39)

(40)

(41)

(42)

(43)

(44) (45) (46) (47)

Persikov, A. V., Ramshaw, J. A. M., Kirkpatrick, A., and Brodsky, B. (2005) Electrostatic interactions involving lysine make major contributions to collagen triplehelix stability. Biochemistry 44, 1414-1422. Fallas, J. A., Dong, J. H., Tao, Y. Z. J., and Hartgerink, J. D. (2012) Structural insights into charge pair interactions in triple helical collagen-like proteins. J. Biol. Chem. 287, 8039-8047. Fallas, J. A., O'Leary, L. E. R., and Hartgerink, J. D. (2010) Synthetic collagen mimics: self-assembly of homotrimers, heterotrimers and higher order structures. Chem. Soc. Rev. 39, 3510-3527. Koide, T., Yuguchi, M., Kawakita, M., and Konno, H. (2002) Metal-assisted stabilization and probing of collagenous triple helices. J. Am. Chem. Soc. 124, 9388-9389. Parmar, A. S., Xu, F., Pike, D. H., Belure, S. V., Hasan, N. F., Drzewiecki, K. E., Shreiber, D. I., and Nanda, V. (2015) Metal stabilization of collagen and de novo designed mimetic peptides. Biochemistry 54, 4987-4997. Chung, E., Keele, E. M., and Miller, E. J. (1974) Isolation and characterization of cyanogen-bromide peptides from α1(III) chain of human collagen. Biochemistry 13, 3459-3464. Bachinger, H. P., Bruckner, P., Timpl, R., Prockop, D. J., and Engel, J. (1980) Folding mechanism of the triple helix in type-III collagen and type-III pN-collagen. Role of disulfide bridges and peptide-bond isomerization. Eur. J. Biochem. 106, 619-632. Barth, D., Kyrieleis, O., Frank, S., Renner, C., and Moroder, L. (2003) The role of cystine knots in collagen folding and stability, part II. Conformational properties of (ProHyp-Gly)(n) model trimers with N- and C-terminal collagen type III cystine knots. Chem. Eur. J. 9, 3703-3714. Mechling, D. E., and Bachinger, H. P. (2000) The collagen-like peptide (GER)15GPCCG forms pH-dependent covalently linked triple helical trimers. J. Biol. Chem. 275, 1453214536. Krishna, O. D., and Kiick, K. L. (2009) Supramolecular assembly of electrostatically stabilized, hydroxyproline-lacking collagen-mimetic peptides. Biomacromolecules 10, 2626-2631. Feng, Y. B., Melacini, G., Taulane, J. P., and Goodman, M. (1996) Acetyl-terminated and template-assembled collagen-based polypeptides composed of Gly-Pro-Hyp sequences. 2. Synthesis and conformational analysis by circular dichroism, ultraviolet absorbance, and optical rotation. J. Am. Chem. Soc. 118, 10351-10358. Kwak, J., De Capua, A., Locardi, E., and Goodman, M. (2002) TREN (tris(2aminoethyl)amine): An effective scaffold for the assembly of triple helical collagen mimetic structures. J. Am. Chem. Soc. 124, 14085-14091. Frank, S., Kammerer, R. A., Mechling, D., Schulthess, T., Landwehr, R., Bann, J., Guo, Y., Lustig, A., Bachinger, H. P., and Engel, J. (2001) Stabilization of short collagen-like triple helices by protein engineering. J. Mol. Biol. 308, 1081-1089. Fields, G. B., Lauer, J. L., Dori, Y., Forns, P., Yu, Y. C., and Tirrell, M. (1998) Proteinlike molecular architecture: Biomaterial applications for inducing cellular receptor binding and signal transduction. Biopolymers 47, 143-151. Yu, Y. C., Berndt, P., Tirrell, M., and Fields, G. B. (1996) Self-assembling amphiphiles for construction of protein molecular architecture. J. Am. Chem. Soc. 118, 12515-12520. Gore, T., Dori, Y., Talmon, Y., Tirrell, M., and Bianco-Peled, H. (2001) Self-assembly of model collagen peptide amphiphiles. Langmuir 17, 5352-5360. Luo, T. Z., and Kiick, K. L. (2013) Collagen-like peptides and peptide-polymer conjugates in the design of assembled materials. Eur. Polym. J. 49, 2998-3009. Newberry, R. W., VanVeller, B., and Raines, R. T. (2015) Thioamides in the collagen triple helix. Chem. Commun. 51, 9624-9627.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(48) (49) (50) (51) (52)

(53) (54) (55) (56)

(57)

(58)

(59) (60)

(61)

(62) (63)

(64)

(65)

(66)

(67)

Zhang, Y. T., Malamakal, R. M., and Chenoweth, D. M. (2015) Aza-glycine induces collagen hyperstability. J. Am. Chem. Soc. 137, 12422-12425. Zhang, Y. T., Herling, M., and Chenoweth, D. M. (2016) General solution for stabilizing triple helical collagen. J. Am. Chem. Soc. 138, 9751-9754. Proulx, C., Sabatino, D., Hopewell, R., Spiegel, J., Ramos, Y. G., and Lubell, W. D. (2011) Azapeptides and their therapeutic potential. Future Med. Chem. 3, 1139-1164. Zega, A. (2005) Azapeptides as pharmacological agents. Curr. Med. Chem. 12, 589-597. Kar, K., Amin, P., Bryan, M. A., Persikov, A. V., Mohs, A., Wang, Y. H., and Brodsky, B. (2006) Self-association of collagen triple helix peptides into higher order structures. J. Biol. Chem. 281, 33283-33290. Gottlieb, D., Morin, S. A., Jin, S., and Raines, R. T. (2008) Self-assembled collagen-like peptide fibers as templates for metallic nanowires. J. Mater. Chem. 18, 3865-3870. Yamazaki, C. M., Asada, S., Kitagawa, K., and Koide, T. (2008) Artificial collagen gels via self-assembly of de novo designed peptides. Biopolymers 90, 816-823. Kotch, F. W., and Raines, R. T. (2006) Self-assembly of synthetic collagen triple helices. P. Natl. Acad. Sci. U.S.A. 103, 3028-3033. Koide, T., Homma, D. L., Asada, S., and Kitagawa, K. (2005) Self-complementary peptides for the formation of collagen-like triple helical supramolecules. Bioorg. Med. Chem. Lett. 15, 5230-5233. Chen, C. C., Hsu, W., Kao, T. C., and Horng, J. C. (2011) Self-assembly of short collagen-related peptides into fibrils via cation-π interactions. Biochemistry 50, 23812383. Kar, K., Ibrar, S., Nanda, V., Getz, T. M., Kunapuli, S. P., and Brodsky, B. (2009) Aromatic interactions promote self-association of collagen triple-helical peptides to higher-order structures. Biochemistry 48, 7959-7968. Rele, S., Song, Y. H., Apkarian, R. P., Qu, Z., Conticello, V. P., and Chaikof, E. L. (2007) D-periodic collagen-mimetic microfibers. J. Am. Chem. Soc. 129, 14780-14787. O'Leary, L. E. R., Fallas, J. A., Bakota, E. L., Kang, M. K., and Hartgerink, J. D. (2011) Multi-hierarchical self-assembly of a collagen mimetic peptide from triple helix to nanofibre and hydrogel. Nat. Chem. 3, 821-828. Jiang, T., Xu, C. F., Zuo, X. B., and Conticello, V. P. (2014) Structurally homogeneous nanosheets from self-assembly of a collagen-mimetic peptide. Angew. Chem. Int. Edit. 53, 8367-8371. He, L. R., and Theato, P. (2013) Collagen and collagen mimetic peptide conjugates in polymer science. Eur. Polym. J. 49, 2986-2997. Zhao, X., Sun, H., Zhang, X. Q., Ren, J., Shao, F., Liu, K., Li, W., and Zhang, A. F. (2016) OEGylated collagen mimetic polypeptides with enhanced supramolecular assembly. Polymer 99, 281-291. Pires, M. M., Przybyla, D. E., Perez, C. M. R., and Chmielewski, J. (2011) Metalmediated tandem coassembly of collagen peptides into banded microstructures. J. Am. Chem. Soc. 133, 14469-14471. He, M. M., Wang, L., Wu, J., and Xiao, J. X. (2016) Ln3+-mediated self-assembly of a collagen peptide into luminescent banded helical nanoropes. Chem. -Eur. J. 22, 19141917. Pires, M. M., Lee, J., Ernenwein, D., and Chmielewski, J. (2012) Controlling the morphology of metal-promoted higher ordered assemblies of collagen peptides with varied core lengths. Langmuir 28, 1993-1997. Przybyla, D. E., and Chmielewski, J. (2008) Metal-triggered radial self-assembly of collagen peptide fibers. J. Am. Chem. Soc. 130, 12610-+.


Page 18 of 23

Page 19 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(68)

(69) (70) (71)

(72)

(73)

(74)

(75)

(76)

(77)

(78) (79)

(80) (81)

(82)

(83)

(84) (85)

Przybyla, D. E., Perez, C. M. R., Gleaton, J., Nandwana, V., and Chmielewski, J. (2013) Hierarchical assembly of collagen peptide triple helices into curved disks and metal ionpromoted hollow spheres. J. Am. Chem. Soc. 135, 3418-3422. Gleaton, J., and Chmielewski, J. (2015) Thermally controlled collagen peptide cages for biopolymer delivery. ACS Biomater.-Sci. Eng. 1, 1002-1008. Wang, A. Y., Mo, X., Chen, C. S., and Yu, S. M. (2005) Facile modification of collagen directed by collagen mimetic peptides. J. Am. Chem. Soc. 127, 4130-4131. Li, Y., Ho, D., Meng, H., Chan, T. R., An, B., Yu, H., Brodsky, B., Jun, A. S., and Yu, S. M. (2013) Direct detection of collagenous proteins by fluorescently labeled collagen mimetic peptides. Bioconjugate Chem. 24, 9-16. Wang, A. Y., Foss, C. A., Leong, S., Mo, X., Pomper, M. G., and Yu, S. M. (2008) Spatio-temporal modification of collagen scaffolds mediated by triple helical propensity. Biomacromolecules 9, 1755-1763. Wang, A. Y., Leong, S., Liang, Y. C., Huang, R. C. C., Chen, C. S., and Yu, S. M. (2008) Immobilization of growth factors on collagen scaffolds mediated by polyanionic collagen mimetic peptides and its effect on endothelial cell morphogenesis. Biomacromolecules 9, 2929-2936. Li, Y., Foss, C. A., Summerfield, D. D., Doyle, J. J., Torok, C. M., Dietz, H. C., Pomper, M. G., and Yu, S. M. (2012) Targeting collagen strands by photo-triggered triple-helix hybridization. P. Natl. Acad. Sci. U.S.A. 109, 14767-14772. Mo, X., An, Y. J., Yun, C. S., and Yu, S. M. (2006) Nanoparticle-assisted visualization of binding interactions between collagen mimetic peptide and collagen fibers. Angew. Chem. Int. Edit. 45, 2267-2270. San, B. H., Li, Y., Tarbet, E. B., and Yu, S. M. (2016) Nanoparticle assembly and gelatin binding mediated by triple helical collagen mimetic peptide. ACS Appl. Mater. Interfaces 8, 19908-19916. Santos, J. L., Li, Y., Culver, H. R., Yu, M. S., and Herrera-Alonso, M. (2014) Conducting polymer nanoparticles decorated with collagen mimetic peptides for collagen targeting. Chem. Commun. 50, 15045-15048. Urello, M. A., Kiick, K. L., and Sullivan, M. O. (2014) A CMP-based method for tunable, cell-mediated gene delivery from collagen scaffolds. J. Mat. Chem. B 2, 8174-8185. Urello, M. A., Kiick, K. L., and Sullivan, M. O. (2016) Integration of growth factor gene delivery with collagen-triggered wound repair cascades using collagen-mimetic peptides. Bioengineering and Translational Medicine 2, 207-219. Perez, C. M. R., Rank, L. A., and Chmielewski, J. (2014) Tuning the thermosensitive properties of hybrid collagen peptide-polymer hydrogels. Chem. Commun. 50, 8174-8176. Islam, M. M., Ravichandran, R., Olsen, D., Ljunggren, M. K., Fagerholm, P., Lee, C. J., Griffith, M., and Phopase, J. (2016) Self-assembled collagen-like-peptide implants as alternatives to human donor corneal transplantation. RSC Adv. 6, 55745-55749. Lee, H. J., Lee, J. S., Chansakul, T., Yu, C., Elisseeff, J. H., and Yu, S. M. (2006) Collagen mimetic peptide-conjugated photopolymerizable PEG hydrogel. Biomaterials 27, 5268-5276. Lee, H. J., Yu, C., Chansakul, T., Hwang, N. S., Varghese, S., Yu, S. M., and Elisseeff, J. H. (2008) Enhanced chondrogenesis of mesenchymal stem cells in collagen mimetic peptide-mediated microenvironment. Tissue Eng. Part A 14, 1843-1851. Luo, J. N., and Tong, Y. W. (2011) Self-assembly of collagen-mimetic peptide amphiphiles into biofunctional nanofiber. Acs Nano 5, 7739-7747. Krishna, O. D., Jha, A. K., Jia, X. Q., and Kiick, K. L. (2011) Integrin-mediated adhesion and proliferation of human MSCs elicited by a hydroxyproline-lacking, collagen-like peptide. Biomaterials 32, 6412-6424.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(86)

(87)

(88)

(89)

(90)

(91)

(92) (93) (94) (95) (96) (97) (98)

(99)

(100)

(101) (102)

(103)

(104)

Hernandez-Gordillo, V., and Chmielewski, J. (2014) Mimicking the extracellular matrix with functionalized, metal-assembled collagen peptide scaffolds. Biomaterials 35, 73637373. Zhang, Z., Lai, Y. X., Yu, L., and Ding, J. D. (2010) Effects of immobilizing sites of RGD peptides in amphiphilic block copolymers on efficacy of cell adhesion. Biomaterials 31, 7873-7882. Humtsoe, J. O., Kim, J. K., Xu, Y., Keene, D. R., Hook, M., Lukomski, S., and Wary, K. K. (2005) A streptococcal collagen-like protein interacts with the α2β1 integrin and induces intracellular signaling. J. Biol. Chem. 280, 13848-13857. Pechar, M., Brus, J., Kostka, L., Konak, C., Urbanova, M., and Slouf, M. (2007) Thermoresponsive self-assembly of short elastin-like polypentapeptides and their poly(ethylene glycol) derivatives. Macromol. Biosci. 7, 56-69. Vandermeulen, G. W. M., Tziatzios, C., and Klok, H. A. (2003) Reversible selforganization of poly(ethylene glycol)-based hybrid block copolymers mediated by a de novo four-stranded α-helical coiled coil motif. Macromolecules 36, 4107-4114. Rosler, A., Klok, H. A., Hamley, I. W., Castelletto, V., and Mykhaylyk, O. O. (2003) Nanoscale structure of poly(ethylene glycol) hybrid block copolymers containing amphiphilic β-strand peptide sequences. Biomacromolecules 4, 859-863. Top, A., Roberts, C. J., and Kiick, K. L. (2011) Conformational and aggregation properties of a PEGylated alanine-rich polypeptide. Biomacromolecules 12, 2184-2192. Top, A. (2011) Controlling assembly of helical peptides via PEGylation strategies. Soft Matter. Roberts, M. J., Bentley, M. D., and Harris, J. M. (2002) Chemistry for peptide and protein PEGylation. Adv. Drug Deliver. Rev. 54, 459-476. Kinstler, O., Molineux, G., Treuheit, M., Ladd, D., and Gegg, C. (2002) Mono-Nterminal poly(ethylene glycol)-protein conjugates. Adv. Drug Deliver. Rev. 54, 477-485. Nilsson, B. L., Kiessling, L. L., and Raines, R. T. (2001) High-yielding Staudinger ligation of a phosphinothioester and azide to form a peptide. Org Lett 3, 9-12. Huisgen, R. (1963) 1.3-Dipolare cycloadditionen - ruckschau und ausblick. Angew. Chem. Int. Edit. 75, 604-+. Tornoe, C. W., Christensen, C., and Meldal, M. (2002) Peptidotriazoles on solid phase: 1,2,3 -triazoles by regiospecific copper(I)-catalyzed 1,3-dipolar cycloadditions of terminal alkynes to azides. J. Org. Chem. 67, 3057-3064. Rostovtsev, V. V., Green, L. G., Fokin, V. V., and Sharpless, K. B. (2002) A stepwise Huisgen cycloaddition process: Copper(I)-catalyzed regioselective "ligation" of azides and terminal alkynes. Angew. Chem. Int. Edit. 41, 2596-+. Dirks, A. J. T., van Berkel, S. S., Hatzakis, N. S., Opsteen, J. A., van Delft, F. L., Cornelissen, J., Rowan, A. E., van Hest, J. C. M., Rutjes, F., and Nolte, R. J. M. (2005) Preparation of biohybrid amphiphiles via the copper catalysed Huisgen 3+2 dipolar cycloaddition reaction. Chem. Commun., 4172-4174. Lowe, A. B. (2010) Thiol-ene "click" reactions and recent applications in polymer and materials synthesis. Polym. Chem. 1, 17-36. Mather, B. D., Viswanathan, K., Miller, K. M., and Long, T. E. (2006) Michael addition reactions in macromolecular design for emerging technologies. Prog. Polym. Sci. 31, 487-531. Friedman, M., Cavins, J. F., and Wall, J. S. (1965) Relative nucleophilic reactivities of amino groups and mercaptide ions in addition reactions with α,β-unsaturated compounds. J. Am. Chem. Soc. 87, 3672-&. Baldwin, A. D., and Kiick, K. L. (2011) Tunable degradation of maleimide-thiol adducts in reducing environments. Bioconjugate Chem. 22, 1946-1953.


Page 20 of 23

Page 21 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(105) (106) (107)

(108)

(109) (110) (111) (112) (113)

(114) (115)

(116) (117)

(118)

(119) (120) (121) (122) (123) (124) (125) (126)

Carlsen, A., and Lecommandoux, S. (2009) Self-assembly of polypeptide-based block copolymer amphiphiles. Curr. Opin. Colloid Interface Sci. 14, 329-339. Chow, D. (2008) Peptide-based biopolymers in biomedicine and biotechnology. Ding, J. X., Xiao, C. S., Tang, Z. H., Zhuang, X. L., and Chen, X. S. (2011) Highly efficient "grafting from" an α-helical polypeptide backbone by atom transfer radical polymerization. Macromol. Biosci. 11, 192-198. Wigenius, J., Bjork, P., Hamedi, M., and Aili, D. (2010) Supramolecular assembly of designed α-helical polypeptide-based nanostructures and luminescent conjugated polyelectrolytes. Macromol. Biosci. 10, 836-841. Shu, J. Y., Tan, C., DeGrado, W. F., and Xu, T. (2008) New design of helix bundle peptide-polymer conjugates. Biomacromolecules 9, 2111-2117. Sahin, E., and Kiick, K. L. (2009) Macromolecule-induced assembly of coiled-coils in alternating multiblock polymers. Biomacromolecules 10, 2740-2749. Mondal, S., and Gazit, E. (2016) The self-assembly of helical peptide building blocks. ChemNanoMat 2, 323-332. Kopecek, J., and Yang, J. Y. (2011) Self-assembly of hybrid copolymers grafted with βsheet and α-helix peptides. Abstr. Pap. Am. Chem. Soc. 241. Elder, A. N., Dangelo, N. M., Kim, S. C., and Washburn, N. R. (2011) Conjugation of βsheet peptides to modify the rheological properties of hyaluronic acid. Biomacromolecules 12, 2610-2616. Lee, O. S., Liu, Y. M., and Schatz, G. C. (2012) Molecular dynamics simulation of βsheet formation in self-assembled peptide amphiphile fibers. J. Nanopart. Res. 14. Kim, S., Kim, J. H., Lee, J. S., and Park, C. B. (2015) Βeta-sheet-forming, self-assembled peptide nanomaterials towards optical, energy, and healthcare applications. Small 11, 3623-3640. Aluri, S., Pastuszka, M. K., Moses, A. S., and MacKay, J. A. (2012) Elastin-like peptide amphiphiles form nanofibers with tunable length. Biomacromolecules 13, 2645-2654. Grieshaber, S. E., Farran, A. J. E., Lin-Gibson, S., Kiick, K. L., and Jia, X. Q. (2009) Synthesis and characterization of elastin-mimetic hybrid polymers with multiblock, alternating molecular architecture and elastomeric properties. Macromolecules 42, 25322541. Grieshaber, S. E., Farran, A. J. E., Bai, S., Kiick, K. L., and Jia, X. Q. (2012) Tuning the properties of elastin mimetic hybrid copolymers via a modular polymerization method. Biomacromolecules 13, 1774-1786. Rodriguez-Cabello, J. C., Arias, F. J., Rodrigo, M. A., and Girotti, A. (2016) Elastin-like polypeptides in drug delivery. Adv. Drug Deliver. Rev. 97, 85-100. Navon, Y., and Bitton, R. (2016) Elastin-like peptides (ELPs) - building blocks for stimuli-responsive self-assembled materials. Isr. J. Chem. 56, 581-589. Saxena, R., and Nanjan, M. J. (2015) Elastin-like polypeptides and their applications in anticancer drug delivery systems: a review. Drug Deliv. 22, 156-167. MacEwan, S. R., and Chilkoti, A. (2014) Applications of elastin-like polypeptides in drug delivery. J. Control Release 190, 314-330. Foster, J. A., Bruenger, E., Gray, W. R., and Sandberg, L. B. (1973) Isolation and aminoacid sequences of tropoelastin peptides. J. Biol. Chem. 248, 2876-2879. Li, B., Alonso, D. O. V., and Daggett, V. (2001) The molecular basis for the inverse temperature transition of elastin. J. Mol. Biol. 305, 581-592. Kojima, C., and Irie, K. (2013) Synthesis of temperature-dependent elastin-like peptidemodified dendrimer for drug delivery. Biopolymers 100, 714-21. Massodi, I., Bidwell, G. L., and Raucher, D. (2005) Evaluation of cell penetrating peptides fused to elastin-like polypeptide for drug delivery. J. Control Release 108, 396408.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

(127)

(128) (129)

(130)

(131)

(132)

(133) (134)

(135)

(136)

(137)

(138)

(139)

(140)

(141) (142)

(143)

Hassouneh, W., Zhulina, E. B., Chilkoti, A., and Rubinstein, M. (2015) Elastin-like polypeptide diblock copolymers self-assemble into weak micelles. Macromolecules 48, 4183-4195. MacEwan, S. R., and Chilkoti, A. (2014) Controlled apoptosis by a thermally toggled nanoscale amplifier of cellular uptake. Nano Lett. 14, 2058-2064. Shamji, M. F., Chen, J., Friedman, A. H., Richardson, W. J., Chilkoti, A., and Setton, L. A. (2008) Synthesis and characterization of a thermally-responsive tumor necrosis factor antagonist. J. Control Release 129, 179-186. McDaniel, J. R., Bhattacharyya, J., Vargo, K. B., Hassouneh, W., Hammer, D. A., and Chilkoti, A. (2013) Self-assembly of thermally responsive nanoparticles of a genetically encoded peptide polymer by drug conjugation. Angew. Chem. Int. Edit. 52, 1683-1687. Callahan, D. J., Liu, W. E., Li, X. H., Dreher, M. R., Hassouneh, W., Kim, M., Marszalek, P., and Chilkoti, A. (2012) Triple stimulus-responsive polypeptide nanoparticles that enhance intratumoral spatial distribution. Nano Lett. 12, 2165-2170. Wright, E. R., McMillan, R. A., Cooper, A., Apkarian, R. P., and Conticello, V. P. (2002) Thermoplastic elastomer hydrogels via self-assembly of an elastin-mimetic triblock polypeptide. Adv. Funct. Mater. 12, 149-154. Wright, E. R., and Conticello, V. P. (2002) Self-assembly of block copolymers derived from elastin-mimetic polypeptide sequences. Adv. Drug Deliver. Rev. 54, 1057-1073. Wright, E. R., Conticello, V. P., and Apkarian, R. P. (2003) Morphological characterization of elastin-mimetic block copolymers utilizing cryo- and cryoetchHRSEM. Microsc. microanal. 9, 171-182. Lee, T. A. T., Cooper, A., Apkarian, R. P., and Conticello, V. P. (2000) Thermoreversible self-assembly of nanoparticles derived from elastin-mimetic polypeptides. Adv. Mater. 12, 1105-+. Haghpanah, J. S., Tu, R., Da Silva, S., Yan, D., Mueller, S., Weder, C., Foster, E. J., Sacui, I., Gilman, J. W., and Montclare, J. K. (2013) Bionanocomposites: differential effects of cellulose nanocrystals on protein diblock copolymers. Biomacromolecules 14, 4360-4367. Yuvienco, C., More, H. T., Haghpanah, J. S., Tu, R. S., and Montclare, J. K. (2012) Modulating supramolecular assemblies and mechanical properties of engineered protein materials by fluorinated amino acids. Biomacromolecules 13, 2273-2278. Dai, M., Haghpanah, J., Singh, N., Roth, E. W., Liang, A., Tu, R. S., and Montclare, J. K. (2011) Artificial protein block polymer libraries bearing two SADs: Effects of elastin domain repeats. Biomacromolecules 12, 4240-4246. Haghpanah, J. S., Yuvienco, C., Roth, E. W., Liang, A., Tu, R. S., and Montclare, J. K. (2010) Supramolecular assembly and small molecule recognition by genetically engineered protein block polymers composed of two SADs. Molecular Biosystems 6, 1662-1667. Haghpanah, J. S., Yuvienco, C., Civay, D. E., Barra, H., Baker, P. J., Khapli, S., Voloshchuk, N., Gunasekar, S. K., Muthukumar, M., and Montclare, J. K. (2009) Artificial protein block copolymers blocks comprising two distinct self-assembling domains. Chembiochem 10, 2733-2735. Cai, L., Dinh, C. B., and Heilshorn, S. C. (2014) One-pot synthesis of elastin-like polypeptide hydrogels with grafted VEGF-mimetic peptides. Biomater. Sci. 2, 757-765. van Eldijk, M. B., Smits, F. C. M., Vermue, N., Debets, M. F., Schoffelen, S., and van Hest, J. C. M. (2014) Synthesis and self-assembly of well-defined elastin-like polypeptide-poly(ethylene glycol) conjugates. Biomacromolecules 15, 2751-2759. Pastuszk, M. K., Wang, X. D., Lock, L. L., Janib, S. M., Cui, H. G., DeLeve, L. D., and MacKay, J. A. (2014) An amphipathic alpha-helical peptide from apolipoprotein A1 stabilizes protein polymer vesicles. J. Control Release 191, 15-23.


Page 22 of 23

Page 23 of 23

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


(144)

(145) (146)

(147)

(148) (149)

(150)

(151) (152)

(153)

Garcia-Arevalo, C., Bermejo-Martin, J. F., Rico, L., Iglesias, V., Martin, L., RodriguezCabello, J. C., and Arias, F. J. (2013) Immunomodulatory nanoparticles from elastin-like recombinamers: single-molecules for tuberculosis vaccine development. Mol. Pharm. 10, 586-597. Nuhn, H., and Klok, H. A. (2008) Secondary structure formation and LCST behavior of short elastin-like peptides. Biomacromolecules 9, 2755-2763. McDaniel, J. R., Radford, D. C., and Chilkoti, A. (2013) A unified model for de novo design of elastin-like polypeptides with tunable inverse transition temperatures. Biomacromolecules 14, 2866-2872. Rowley, M. J., Williamson, D. J., and Mackay, I. R. (1987) Evidence for local synthesis of antibodies to denatured collagen in the synovium in rheumatoid-arthritis. Arthritis Rheum. 30, 1420-1425. Dodge, G. R., Pidoux, I., and Poole, A. R. (1991) The degradation of type-II collagen in rheumatoid-arthritis - an immunoelectron microscopic study. Matrix 11, 330-338. Van Den Steen, P. E., Proost, P., Grillet, B., Brand, D. D., Kang, A. H., Van Damme, J., and Opdenakker, G. (2002) Cleavage of denatured natural collagen type II by neutrophil gelatinase B reveals enzyme specificity, post-translational modifications in the substrate, and the formation of remnant epitopes in rheumatoid arthritis. FASEB J. 16, 11. Poole, A. R., Kobayashi, M., Yasuda, T., Laverty, S., Mwale, F., Kojima, T., Sakai, T., Wahl, C., El-Maadawy, S., Webb, G., Tchetina, E., and Wu, W. (2002) Type II collagen degradation and its regulation in articular cartilage in osteoarthritis. Ann. Rheum. Dis. 61, 78-81. Loeser, R. F., Goldring, S. R., Scanzello, C. R., and Goldring, M. B. (2012) Osteoarthritis: A disease of the joint as an organ. Arthritis Rheum. 64, 1697-1707. Lindeman, J. H. N., Ashcroft, B. A., Beenakker, J. W. M., van Es, M., Koekkoek, N. B. R., Prins, F. A., Tielemans, J. F., Abdul-Hussien, H., Bank, R. A., and Oosterkamp, T. H. (2010) Distinct defects in collagen microarchitecture underlie vessel-wall failure in advanced abdominal aneurysms and aneurysms in Marfan syndrome. P. Natl. Acad. Sci. U.S.A. 107, 862-865. Kuznetsova, N. V., McBride, D. J., and Leikin, S. (2003) Changes in thermal stability and microunfolding pattern of collagen helix resulting from the loss of α2(I) chain in osteogenesis imperfecta murine. J. Mol. Biol. 331, 191-200.


Collagen-like peptide bioconjugates - Bioconjugate Chemistry (ACS

Recommend Documents