Letters Cite This: ACS Chem. Biol. 2017, 12, 2720-2724
pubs.acs.org/acschemicalbiology
Selective Targeting of Vibrios by Fluorescent Siderophore-Based Probes Peng-Hsun Chase Chen,† Sheng-Yang Ho,†,§ Pin-Lung Chen,†,§ Tzu-Chiao Hung,‡,§ An-Jou Liang,‡ Tang-Feng Kuo,† Hsiao-Chun Huang,‡ and Tsung-Shing Andrew Wang*,† †
Department of Chemistry, National Taiwan University, Taipei, 10617, Taiwan (Republic of China) Institute of Molecular and Cellular Biology, National Taiwan University, Taipei, 10617, Taiwan (Republic of China)
‡
S Supporting Information *
ABSTRACT: Siderophores are small molecules used to specifically transport iron into bacteria via related receptors. By adapting siderophores and hijacking their pathways, we may discover an efficient and selective way to target microbes. Herein, we report the synthesis of a siderophore-fluorophore conjugate VF-FL derived from vibrioferrin (VF). Using flow cytometry and fluorescence microscopy, the probe selectively labeled vibrios, including V. parahaemolyticus, V. cholerae, and V. vulnif icus, even in the presence of other species such as S. aureus and E. coli. The labeling is siderophore-related and both iron-limited conditions and the siderophore moiety are required. The competitive relationship between VF-FL and VF in vibrios implies an unreported VF-related transport mechanism in V. cholerae and V. vulnificus. These studies demonstrate that the siderophore scaffold provides a method to selectively target microbes expressing cognate receptors under iron-limited conditions.
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bacteria produce these high-affinity iron chelators to scavenge all possible iron from their surroundings. Upon binding to iron, the complexes are recognized by their cognate receptors and transported into bacteria.19 Thus, by harnessing the siderophore and hijacking this pathway, this iron acquisition mechanism can be adapted into various applications. In several previous studies, siderophore-based strategies have been developed into detecting probes,20−24 pathogen capturing materials,25 or drug delivery agents.26−30 Nevertheless, a key challenge remaining is the improvement of strain selectivity since several siderophores, including entereobactin, desferrixamine B, or ferrichrome, can be utilized by a wide spectrum of microbes.31 On the other hand, while other conjugates, such as those based on pyochelin, 21 demonstrate better strain specificity, they exhibit poor labeling efficiency. Consequently, to further expand the toolbox of siderophore conjugates, increased specificity and high labeling efficiency are required. While searching for the candidate, we focused on the vibrio species. These Gram-negative, halophilic bacteria have been categorized as common pathogens that could cause cholera, septicemia, gastroenteritis, etc.32,33 Moreover, as they are abundant aquatic microbes and part of aquatic microbiota, many vibrios form symbiotic relationships with marine animals and are frequently blamed for foodborne illnesses.34 Consid-
here is mounting evidence that the bacterial community can affect the physiology of hosts. For example, it has been suggested that human gut microbiota play roles in modulating host metabolism,1,2 immunity,3 neural communication,4 and pathogenic bacterial activity.5−7 Thus, understanding how bacteria interact with other bacteria and their hosts provides not only fundamental advances in biology but also underlying applications for human healthcare management. Bacterial identification is critical in the field of microbiota research, and many current techniques, including 16S rRNA sequencing 8,9 and fluorescence in situ hybridization (FISH),10,11 have shed light on the composition of microbiota samples. However, these techniques are highly technical, often requiring laborious sample preparation, and provide only limited temporal or spatial information. Other approaches such as those based on antibodies,12 antibiotics,13 antimicrobial peptides14 or sugars,15,16 results in poor bacterial selectivity when complex bacterial samples are analyzed. Therefore, new bacterial targeting probes with facile handling and improved species-selectivity are in great demand. The symbiotic interaction between bacteria and other species results in highly diverse biological networks and the evolution of bioactive secondary metabolites, which represent an excellent source of structural skeletons for drug development. One particular type of secondary metabolite, siderophores, was developed to acquire iron. Siderophores are iron chelators secreted and utilized by many microbes.17,18 Under ironlimiting conditions, such as inside vertebrate hosts, many © 2017 American Chemical Society
Received: August 4, 2017 Accepted: October 9, 2017 Published: October 9, 2017 2720
DOI: 10.1021/acschembio.7b00667 ACS Chem. Biol. 2017, 12, 2720−2724
Letters
ACS Chemical Biology Scheme 1. Synthesis of Vibrios Targeting Probe VF-FL
not only V. parahaemolyticus but also V. cholerae and V. vulnif icus gave significant fluorescence increases, while all other bacteria were irresponsive under iron-limited conditions (Figure 1a). To test if these results were attributable to the iron-dependent acquisition pathway, we examined different iron conditions. After incubation in minimal media containing 170 μM of iron(III) to down-regulate the uptake pathway, none of them showed fluorescent enhancement when treated with VFFL, a result that agreed with our hypothesis (Figure 1b). Since no known VF receptors have been identified in V. cholerae and V. vulnificus, it is not clear how these two strains can acquire VF. Nevertheless, further experiments also demonstrated clear dose- and time-dependent fluorescent responses of these vibrios with VF-FL (Supporting Information Figures S6a,b, S7a,b). Thus, to confirm that the VF-FL was transported through the VF-dependent mechanism, we performed a competition assay, whereby excess VF was added to compete with VF-FL uptake. Using V. parahaemolyticus as a positive control, a 2.6 fold fluorescence increase is detected with 5 μM VF-FL, while the cotreatment with exogenous VF resulted in a dose-dependent decrease in fluorescence. The same competition performed in V. cholerae and V. vulnificus showed a clear dose-dependent fluorescence decrease in both cases, suggesting a possible unknown VFdependent mechanism in those two strains (Figure 1c). The ability of VF-FL to target vibrios was also investigated via epifluorescence microscopy. When V. parahaemolyticus, V. cholerae, or V. vulnificus was treated with VF-FL under ironlimited conditions, all were labeled with high efficiency (Figure 2a, Supporting Information Figure S8), whereas no fluorescent labeling was observed with azFL. By comparison, when the same labeling protocol was applied on S. aureus and E. coli, neither of them showed a response to VF-FL or azFL (Figure 2b). The strain selectivity of VF-FL was also demonstrated in the mixed bacteria samples, in which V. parahaemolyticus, V. cholerae, or V. vulnif icus was coincubated with S. aureus or E. coli and treated with VF-FL under iron-limited conditions. The fluorescent images clearly showed that only vibrios (curved-rod shape) were selectively labeled, but not S. aureus (round shape) or E. coli (rod shape; Figure 3a, Supporting Information Figures S9a, S10a). Compared to S. aureus and E. coli, since vibrios showed different FSC-SSC (forward scatter-side scatter) patterns in flow cytometry (Figure 3b, Supporting Information Figure S14b), it is possible to distinguish vibrios from other strains of bacteria by scatter plots. In contrast to E. coli, S.
ering their high relevance to both human microbiota and aquatic ecology, it is worthwhile to develop probes that target vibrio species. Vibrioferrin (VF), a unique siderophore produced by V. parahaemolyticus and V. alginolyticus,35is exclusive to vibrios (Scheme 1). So far, no reports have shown that bacteria from other genera can express VF receptors, hence VF should be a strain-specific targeting moiety. To demonstrate the feasibility of our design, a fluorophore is attached to assess strain specificity. When developing siderophore conjugates, the introduction of nonnative scaffolds may compromise iron binding or receptor recognition. Due to a lack of crystallographic information, we functionalized VF from the methyl moiety to minimize perturbations. We replaced the L-alanine in VF with L-lysine to facilitate an extendable analogue. The synthesis started with the Boc-Lys(Z)-OH and continued according to a procedure reported by Takeuchi et al.,36 where 2-aminoethanol, chiral (S)dibenzylcitrate 2, and benzyl ketogutaric acid 4 were subsequently linked to afford the protected VF-lysine analogue 6 (or VF*) in comparable yields (Scheme 1). In the final global deprotection, triethylsilane was used for transfer of hydrogenation to provide improved results. An NHS-activated alkyne linker 7 was introduced onto the amino group of VF* selectively to produce the alkyne-functionalized VF (VFalkyne). The azido-modified fluorescein 9 (or azFL) was conjugated with VF-alkyne via copper(I)-catalyzed alkyneazide cycloaddition (CuAAC) to afford final product 10 (or VF-FL). Many synthetic VF analogues demonstrated characteristic features similar to previous reports of native VF, such as (i) the pyrrolidinone ring formation,35 (ii) boron-binding,37 and (iii) photolysis in the presence of iron38 (Supporting Information Figures S1−S3). With VF-FL in hand, we first evaluated the fluorescent response to V. parahaemolyticus, a strain bearing the VF receptor. Upon treatment of VF-FL in the iron-limited minimal media, a dose-dependent fluorescent response was observed after 4 h of incubation at 37 °C by flow cytometry. In contrast, the treatment with azFL under the same conditions showed no significant change (Supporting Information Figure S5b). This suggests that VF is responsible for the fluorescence enhancement. In addition, a clear time dependent fluorescence enhancement was also observed (Supporting Information Figure S5a). To assess the specificity of our probe, V. parahaemolyticus along with S. aureus, B. subtilis, E. coli, S. enterica, V. cholerae, and V. vulnificus were tested. Surprisingly, 2721
DOI: 10.1021/acschembio.7b00667 ACS Chem. Biol. 2017, 12, 2720−2724
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ACS Chemical Biology
Figure 2. Epifluorescence microscopy images. (a) V. parahaemolyticus (VP) treated with azFL (top) or VF-FL (bottom). (b) E. coli (EC, top) and S. aureus (SA, bottom) treated with VF-FL. Scale bar: 10 μm (full view) or 1 μm (enlarged view). All bacteria tested were precultured in iron-limited minimal media. Data were acquired from samples treated with 25 μM VF-FL at 37 °C for 4 h. Figure 1. Fluorescence response of the VF-FL uptake by bacteria, S. aureus (SA), B. subtilis (BS), E. coli (EC), S. enterica (SE), V. cholerae (VC), V. vulnif icus (VV), and V. parahaemolyticus (VP), precultured in (a) iron-limited (Fe3+: 1.7 μM) and (b) iron-rich (Fe3+: 170 μM) TBSM minimal media. Bars represent fluorescence intensity measured by flow cytometry (mean ± SEM, n ≥ 3) with (+) and without (−) VF-FL treatment. Data were acquired from samples treated with 25 μM of VF-FL at 37 °C for 4 h. (c) Fluorescence response of the VFFL uptake by V. parahaemolyticus (VP), V. cholerae (VC), and V. vulnificus (VV) in the presence of exogenous VF. Bars represent fluorescent intensity measured by flow cytometry (mean ± SEM, n ≥ 3). All bacteria tested were precultured in iron-limited minimal media. Data were acquired by samples treated with 5 μM of VF-FL and 10, 25, 50, and 100 equiv of VF at 37 °C for 4 h.
bacteria from other genera. The targeting process was proven to be siderophore-mediated and iron-related through the VF siderophore moiety and under iron-limited conditions. Although there are no reports on the utilization of VF in V. cholerae and V. vulnif icus, our competition results between VFFL and VF have implied the existence of the VF-associated transport pathway(s) in these vibrios. Although the selective vibrio-targeting of VF-FL is evident, how VF-FL achieved the selectivity remains unclear. It might not be necessary to know the detailed uptake process when using VF-fluorophore conjugates as the vibrio-labeling agent. Nevertheless, if developing VF-antibiotic conjugates is desired, further investigations are required to understand: (i) whether our linking strategy allows the VF conjugates only to bind on membrane receptor(s) or to be transported inside cells and (ii) what receptor(s) are responsible for VF uptake in vibrios. The satisfactory strain-targeting specificity and labeling efficiency rendered our VF-FL probe as an intriguing imaging agent for pathogen detection. However, since vibrio infections are not typically deadly and more convenient and cheaper protocols for rapid vibrio detection or diagnosis already exist, developing alternative methods is not urgent. Moreover, since VF-FL labeling relies on expensive florescence-based techniques, such as flow cytometry or fluorescence microscopy, it is not practical to use our probe for diagnostic purposes. Instead, the ability of VF-FL to specifically label vibrios in a complex microbial sample opens up the possibility to study the host−
aureus overlapped less with vibrios on scatter plots and therefore can be easily sorted to demonstrate the strain selectivity of VF-FL. By mapping scatter plots and fluorescence intensity using flow cytometry, the consistent results were also confirmed where only vibrios were selectively labeled (green scattering pattern and high fluorescence peak) in the presence of S. aureus (blue scattering pattern and low fluorescence peak) treated with VF-FL (Figure 3c, Supporting Information Figures S9b, S10b). In conclusion, we have developed a vibrio-specific fluorescent probe, VF-FL, based on the native VF scaffold. The VF-FL probe can selectively label vibrios, including V. parahaemolyticus, V. cholerae, and V. vulnif icus, even in the presence of 2722
DOI: 10.1021/acschembio.7b00667 ACS Chem. Biol. 2017, 12, 2720−2724
Letters
ACS Chemical Biology
our VF-alkyne provides a platform that allows a wide range of azido-functionalized cargos, including near IR dyes, MRI, and PET imaging moieties, to be easily attached via the CuAAC reaction. This synthetic flexibility allows us to tune the reporter unit for different imaging applications in the future. Our targeting strategy requires iron-limited conditions, a factor that appears to limit potential applications at first glance. However, in the host physiological environment, the iron is often sequestered and maintained at a very low level (∼10−18 M). Consequently, this makes it promising for future in vivo applications.
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METHODS
Experimental methods are detailed in the Supporting Information.
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acschembio.7b00667. The synthesis and characterizations of VF-FL, azFL, functionalized VF and other chemicals used in this study; the procedure for bacterial labeling assays; and other experimental details (PDF)
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AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. ORCID
Tsung-Shing Andrew Wang: 0000-0002-4690-3438 Author Contributions §
These authors contributed equally to this work.
Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS We thank the Ministry of Science and Technology, Taiwan, (MOST 104-2119-M-002-021 and 105-2113-M-002-003 to T.S.A.W.) and Center for Emerging Material and Advanced Devices, National Taiwan University (NTU-ERP-106R880210) for financial support.
Figure 3. Demonstration of VF-FL selective labeling. (a) Epifluorescence microscopy images of V. parahaemolyticus (VP) treated with VF-FL in the presence of S. aureus (SA, top row) and E. coli (EC, bottom row). (b) FSC-SSC plots of V. parahaemolyticus (VP), S. aureus (SA), and E. coli (EC) treated with 25 μM VF-FL by flow cytometry. (c) Flow cytometry analysis of V. parahaemolyticus (VP) treated with VF-FL in the presence of S. aureus (SA). Scale bar: 10 μm (full view) or 1 μm (enlarged view). All bacteria tested were precultured in iron-limited minimal media. Data were acquired from samples treated with 25 μM VF-FL at 37 °C for 4 h.
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REFERENCES
(1) Ley, R. E., Turnbaugh, P. J., Klein, S., and Gordon, J. I. (2006) Microbial ecology: human gut microbes associated with obesity. Nature 444, 1022. (2) Zhang, H., DiBaise, J. K., Zuccolo, A., Kudrna, D., Braidotti, M., Yu, Y., Parameswaran, P., Crowell, M. D., Wing, R., Rittmann, B. E., and Krajmalnik-Brown, R. (2009) Human gut microbiota in obesity and after gastric bypass. Proc. Natl. Acad. Sci. U. S. A. 106, 2365−2370. (3) Round, J. L., and Mazmanian, S. K. (2009) The gut microbiota shapes intestinal immune responses during health and disease. Nat. Rev. Immunol. 9, 313−323. (4) Douglas-Escobar, M., Elliott, E., and Neu, J. (2013) Effect of intestinal microbial ecology on the developing brain. JAMA Pediatr. 167, 374−379. (5) Kamada, N., Chen, G. Y., Inohara, N., and Núñez, G. (2013) Control of pathogens and pathobionts by the gut microbiota. Nat. Immunol. 14, 685−690. (6) Sassone-Corsi, M., and Raffatellu, M. (2015) No vacancy: how beneficial microbes cooperate with immunity to provide colonization resistance to pathogens. J. Immunol. 194, 4081−4087.
pathogen interactions. As successfully demonstrated in a recent work achieved by Ferreira et al.,39 siderophore conjugates could be used in theranostics to image bacterial infection in the smallanimal model. This encourages us that our VF-FL might also be able to probe the spatial and temporal interactions of vibrios within microbiota inside hosts, which should provide us more information to better understand the physiology and ecology of certain species of bacteria within complex microbial communities in hosts. For further in vivo biological applications in infected animal models, our current reporter design in VF-FL might not be optimal due to the limited photophysical and photochemical properties of the fluorescein moiety. Nevertheless, the design of 2723
DOI: 10.1021/acschembio.7b00667 ACS Chem. Biol. 2017, 12, 2720−2724
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ACS Chemical Biology (7) Bäumler, A. J., and Sperandio, V. (2016) Interactions between the microbiota and pathogenic bacteria in the gut. Nature 535, 85−93. (8) Olsen, G. J., Lane, D. J., Giovannoni, S. J., Pace, N. R., and Stahl, D. A. (1986) Microbial ecology and evolution: a ribosomal RNA approach. Annu. Rev. Microbiol. 40, 337−365. (9) Shin, J., Lee, S., Go, M.-J., Lee, S. Y., Kim, S. C., Lee, C.-H., and Cho, B.-K. (2016) Analysis of the mouse gut microbiome using fulllength 16S rRNA amplicon sequencing. Sci. Rep. 6, 29681. (10) Moter, A., and Göbel, U. B. (2000) Fluorescence in situ hybridization (FISH) for direct visualization of microorganisms. J. Microbiol. Methods 41, 85−112. (11) Rigottier-Gois, L., Le Bourhis, A.-G., Gramet, G., Rochet, V., and Doré, J. (2003) Fluorescent hybridisation combined with flow cytometry and hybridisation of total RNA to analyse the composition of microbial communities in human faeces using 16S rRNA probes. FEMS Microbiol. Ecol. 43, 237−245. (12) Nakatsuji, T., Chiang, H.-I., Jiang, S. B., Nagarajan, H., Zengler, K., and Gallo, R. L. (2013) The microbiome extends to subepidermal compartments of normal skin. Nat. Commun. 4, 1431. (13) Van Oosten, M., Schäfer, T., Gazendam, J. A., Ohlsen, K., Tsompanidou, E., De Goffau, M. C., Harmsen, H. J., Crane, L. M., Lim, E., Francis, K. P., et al. (2013) Real-time in vivo imaging of invasive-and biomaterial-associated bacterial infections using fluorescently labelled vancomycin. Nat. Commun. 4, 2584. (14) Akram, A. R., Avlonitis, N., Lilienkampf, A., Perez-Lopez, A. M., McDonald, N., Chankeshwara, S. V., Scholefield, E., Haslett, C., Bradley, M., and Dhaliwal, K. (2015) A labelled-ubiquicidin antimicrobial peptide for immediate in situ optical detection of live bacteria in human alveolar lung tissue. Chem. Sci. 6, 6971−6979. (15) Ning, X., Lee, S., Wang, Z., Kim, D., Stubblefield, B., Gilbert, E., and Murthy, N. (2011) Maltodextrin-based imaging probes detect bacteria in vivo with high sensitivity and specificity. Nat. Mater. 10, 602−607. (16) Geva-Zatorsky, N., Alvarez, D., Hudak, J. E., Reading, N. C., Erturk-Hasdemir, D., Dasgupta, S., Von Andrian, U. H., and Kasper, D. L. (2015) In vivo imaging and tracking of host-microbiota interactions via metabolic labeling of gut anaerobic bacteria. Nat. Med. 21, 1091− 1100. (17) Neilands, J. (1995) Siderophores: structure and function of microbial iron transport compounds. J. Biol. Chem. 270, 26723−26726. (18) Boukhalfa, H., and Crumbliss, A. L. (2002) Chemical aspects of siderophore mediated iron transport. BioMetals 15, 325−339. (19) Krewulak, K. D., and Vogel, H. J. (2008) Structural biology of bacterial iron uptake. Biochim. Biophys. Acta, Biomembr. 1778, 1781− 1804. (20) Nudelman, R., Ardon, O., Hadar, Y., Chen, Y., Libman, J., and Shanzer, A. (1998) Modular fluorescent-labeled siderophore analogues. J. Med. Chem. 41, 1671−1678. (21) Noël, S., Guillon, L., Schalk, I. J., and Mislin, G. t. L. (2011) Synthesis of fluorescent probes based on the pyochelin siderophore scaffold. Org. Lett. 13, 844−847. (22) Kim, H. S., Song, W. Y., and Kim, H. J. (2015) Development of a novel fluorescence probe capable of assessing the cytoplasmic entry of siderophore-based conjugates. Org. Biomol. Chem. 13, 73−76. (23) Lee, A. A., Chen, Y. C. S., Ekalestari, E., Ho, S. Y., Hsu, N. S., Kuo, T. F., and Wang, T. S. A. (2016) Facile and versatile chemoenzymatic synthesis of enterobactin analogues and applications in bacterial detection. Angew. Chem., Int. Ed. 55, 12338−12342. (24) Ho, Y.-H., Ho, S.-Y., Hsu, C.-C., Shie, J.-J., and Wang, T.-S. A. (2017) Utilizing an iron (III)-chelation masking strategy to prepare mono-and bis-functionalized aerobactin analogues for targeting pathogenic bacteria. Chem. Commun. 53, 9265−9268. (25) Kim, Y., Lyvers, D. P., Wei, A., Reifenberger, R. G., and Low, P. S. (2012) Label-free detection of a bacterial pathogen using an immobilized siderophore, deferoxamine. Lab Chip 12, 971−976. (26) Miller, M. J., and Malouin, F. (1993) Microbial iron chelators as drug delivery agents: the rational design and synthesis of siderophoredrug conjugates. Acc. Chem. Res. 26, 241−249.
(27) Ghosh, A., Ghosh, M., Niu, C., Malouin, F., Moellmann, U., and Miller, M. J. (1996) Iron transport-mediated drug delivery using mixed-ligand siderophore-β-lactam conjugates. Chem. Biol. 3, 1011− 1019. (28) Wencewicz, T. A., Long, T. E., Möllmann, U., and Miller, M. J. (2013) Trihydroxamate siderophore−fluoroquinolone conjugates are selective sideromycin antibiotics that target Staphylococcus aureus. Bioconjugate Chem. 24, 473−486. (29) Wencewicz, T. A., and Miller, M. J. (2013) Biscatecholate− monohydroxamate mixed ligand siderophore−carbacephalosporin conjugates are selective sideromycin antibiotics that target Acinetobacter baumannii. J. Med. Chem. 56, 4044−4052. (30) Zheng, T., and Nolan, E. M. (2014) Enterobactin-mediated delivery of β-lactam antibiotics enhances antibacterial activity against pathogenic Escherichia coli. J. Am. Chem. Soc. 136, 9677−9691. (31) Thompson, M. G., Corey, B. W., Si, Y., Craft, D. W., and Zurawski, D. V. (2012) Antibacterial activities of iron chelators against common nosocomial pathogens. Antimicrob. Agents Chemother. 56, 5419−5421. (32) Blake, P. A., Merson, M. H., Weaver, R. E., Hollis, D. G., and Heublein, P. C. (1979) Disease caused by a marine vibrio: clinical characteristics and epidemiology. N. Engl. J. Med. 300, 1−5. (33) Tantillo, G., Fontanarosa, M., Di Pinto, A., and Musti, M. (2004) Updated perspectives on emerging vibrios associated with human infections. Lett. Appl. Microbiol. 39, 117−126. (34) Scallan, E., Hoekstra, R. M., Angulo, F. J., Tauxe, R. V., Widdowson, M.-A., Roy, S. L., Jones, J. L., and Griffin, P. M. (2011) Foodborne illness acquired in the United Statesmajor pathogens. Emerging Infect. Dis. 17, 7. (35) Yamamoto, S., Okujo, N., Yoshida, T., Matsuura, S., and Shinoda, S. (1994) Structure and iron transport activity of vibrioferrin, a new siderophore of Vibrio parahaemolyticus. J. Biochem. 115, 868− 874. (36) Takeuchi, Y., Nagao, Y., Toma, K., Yoshikawa, Y., Akiyama, T., Nishioka, H., Abe, H., Harayama, T., and Yamamoto, S. (1999) Synthesis and Siderophore Activity of Vibrioferrin and One of Its Diastereomeric Isomers. Chem. Pharm. Bull. 47, 1284−1287. (37) Amin, S. A., Küpper, F. C., Green, D. H., Harris, W. R., and Carrano, C. J. (2007) Boron binding by a siderophore isolated from marine bacteria associated with the toxic dinoflagellate Gymnodinium catenatum. J. Am. Chem. Soc. 129, 478−479. (38) Amin, S. A., Green, D. H., Küpper, F. C., and Carrano, C. J. (2009) Vibrioferrin, an unusual marine siderophore: iron binding, photochemistry, and biological implications. Inorg. Chem. 48, 11451− 11458. (39) Ferreira, K., Hu, H. Y., Fetz, V., Prochnow, H., Rais, B., Müller, P. P., and Brönstrup, M. (2017) Multivalent siderophore−DOTAM conjugates as theranostics for imaging and treatment of bacterial infections. Angew. Chem., Int. Ed. 56, 8272−8276.
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