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Interactions of Amino-Containing Peptides with Sodium Silicate and Colloidal Silica: A Biomimetic Approach of Silicification Thibaud Coradin, Olivier Durupthy, and Jacques Livage* Laboratoire de Chimie de la Matie` re Condense´ e, CNRS-UMR 7574, Universite´ Pierre et Marie Curie, 4 place Jussieu, F-75252 Paris cedex 05, France Received July 16, 2001. In Final Form: October 24, 2001 The influence of poly(lysine) and poly(arginine) on silicate solutions and silica sols has been investigated. In both cases, solid formation could be observed that appeared to depend on the polymer chain length. Polyelectrolytes induced gelation of diluted sodium silicate solutions. Studies of the silicic acid content using the colorimetric molybdosilicate method suggested that polymers act as gelating agents of silica oligomers via electrostatic interactions that favor condensation. In the case of silica sols, quasi-elastic light scattering measurements indicate that particles are first adsorbed on the polymer chain to form aggregates that then flocculate in the presence of additional peptides. Structural characterizations of the solids obtained in both cases were consistent with the proposed models. These results are discussed in the frame of biogenic silica formation at the protein/silica interface.
1. Introduction In their search of new methods to synthesize highly structured silica, chemists have turned their attention toward biological systems that are able to use a natural source of silicic acid to build carefully designed architectures.1 As a well-known example, the brown algae diatoms have been extensively studied for quite a long time2 and recently re-examined following a biomimetic approach.3-5 These studies mainly focused on the nature and role of the organic matrix used as a template by the cell to form its silica capsule, named frustule.6,7 As a matter of fact, various subsets of proteins can be found that play distinctive roles in the biosilicification process.8 Of particular interest are the polycationic peptides called silaffins.7 They contain lysine and arginine groups which have been suggested to promote the polycondensation of the silicic acid extracted by the cell from seawater and concentrated in a vesicle where biogenic silica is formed.2 Because silaffin proteins exhibit a large diversity of amino acids, it is difficult to determine the influence of each of these amino acids, and of their sequence, on the biosilicification process. We therefore propose a step-bystep approach to this problem, starting from simple model systems whose complexity may then be progressively increased. The first step presented here is based on the possibility to screen each amino acid group separately using homopeptides that bear only one kind of functionality. Using the molybdosilicate method,9 we have recently * Corresponding author. Phone: 33-1-44273365. Fax: 33-144274769. E-mail:
[email protected]. (1) Parkinson, J.; Gordon, R. TIBTECH 1999, 17, 190-196. (2) Simpson, T. L.; Volcani, B. E. Silicon and siliceous structures in biological systems; Springer: New York, 1981. (3) Morse, D. E. TIBTECH 1999, 17, 230-232. (4) Zaremba, C. M.; Stucky, G. D. Curr. Opin. Solid State Mater. Sci. 1996, 1, 425-429. (5) Cha, J. N.; Stucky, G. D.; Morse, D. E.; Deming, T. J. Nature 2000, 403, 289-292. (6) Hecky, R. E.; Mopper, K.; Kilham, P.; Dgens, E. T. Mar. Biol. 1973, 19, 323-328. (7) Kro¨ger, N.; Deutzmann, R.; Sumper, M. Science 1999, 286, 11291132. (8) Perry, C. C.; Keeling-Tucker, T. J. Biol. Inorg. Chem. 2000, 5, 537-550.
shown10 that amino acids interact poorly with an aqueous silica precursor. In sharp contrast, depending on the nature of the chain side-groups, some homopeptides are able to favor silica formation. Particularly, poly(lysine) appeared to induce gelation. It was suggested that the distribution of positively charged NH3+ groups along the peptide chain could bring negatively charged silica oligomers close enough for condensation to occur, providing a first key to the understanding of the silaffins’ role in biosilicification.10 However, very little is known about the nature of the silica species that interact with the cellular material to build the silica network. It has been shown that diatoms were able to capture monomeric silicic acid, but not colloidal silica, to accumulate a silicon “pool”. Within this reservoir, it was postulated that silicon was stocked in the form of silica particles.2 During the silicification process, silicon is extracted from this pool and transported to the vesicle where frustule formation takes place.2 However, the nature of the extracted and transported silicon species (monomeric silica, silicon complexes, colloidal silica) is not known. Therefore, we have investigated more closely the effect of lysine, arginine, and their homopeptides on both silicic acid solutions and silica particle sols. Electrostatic interactions between positively charged peptides and negatively charged silica species favor the formation of silica. This process appears highly dependent on the polymer chain length. Peptides induced the gelation of silicate solutions and the flocculation of silica sols, leading to mesoporous and meso-to-macroporous solids. Considering poly(lysine) and poly(arginine) as simple but adequate analogues of silaffins,7 these results shed new light on the role of these proteins as active agents in the biosilicification processes. 2. Materials and Methods 2.1. Chemicals. Waterglass (27% SiO2, 10% NaOH) from Riedel-de Hae¨n was chosen as the source of silicic acid. Aqueous silicate solutions were preferred to silicon alkoxide precursors (9) Alexander, B. G. J. Am. Chem. Soc. 1953, 75, 5655-5657. (10) Coradin, T.; Livage, J. Colloids Surf., B 2001, 21, 329-336.
10.1021/la011106q CCC: $22.00 © 2002 American Chemical Society Published on Web 02/22/2002
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Chart 1. Chemical Formulas of Amino Acids Used in This Work
currently used for such studies because they correspond to the usual form of soluble silica in nature.5,7 For similar reasons, diluted solutions (10-2 M) of SiO2 were used. Ludox HS-40 was purchased from Aldrich. Sols contained 40 wt % SiO2 stabilized by NaOH (0.4 wt % Na2O). The particle diameter was 12 nm. Surface charge titration measured in a phosphate buffer (pH ) 7.2) by a pH titration procedure already described11 gives 220-240 negative charges (Si-O- surface groups) per particle. L-Lysine (Lys), L-arginine (Arg), poly(L-lysine)n (p-Lys; n ≈ 20, 35, 155, 1005), and poly(L-arginine)n (p-Arg; n ≈ 65) were purchased from Sigma (Chart 1). Molybdosilicate studies were performed in a 0.05 M Tris-HCl buffer (pH ) 7.2). Ammonium heptamolybdate (NH4)6Mo7O24‚4H2O was purchased from Prolabo. Ludox flocculation studies were performed in 0.05 M phosphate buffer (TPO4) (pH ) 7.2). 2.2. Molybdosilicate Titration. In a typical experiment, 65 mg of the sodium silicate solution was diluted in 30 mL of the appropriate buffer in order to obtain a 10-2 M silica solution. The mixture was stirred for 10 min before a solution containing 3 × 10-5 mol of amino acid (or the corresponding quantity of peptide so that the monomer-to-Si ratio was kept equal to 0.1) dissolved in 3 mL of the buffer solution was added. At regular intervals of time, 400 µL portions of the reacting solution were taken and added to 5 mL of deionized water. H2SO4 (200 µL) (1.5 M) and 200 µL of ammonium molybdate (0.08 M) were then added.12 The mixture was left to stand for 10 min in order to allow monomeric silicic acid and small silica oligomers to react with the heptamolybdic acid to form the yellow silicomolybdic acid H8Si(Mo2O7)6. The optical density (OD) of the final solution was measured at 400 nm using a double-beam Kontron Uvikon 940 spectrophotometer. Data were reproducible within a 5% error range. 2.3. Quasi-Elastic Light Scattering (QELS). Average hydrodynamic diameters (DH) were obtained by quasi-elastic light scattering using an AMTEC SM 200 apparatus equipped with a He-Ne laser beam and a Brookhaven Bi2030 correlator. Solutions (1/n mM) of polyelectrolytes were prepared in TPO4 so that the concentration of peptide side chain groups was kept constant to 1 mM. Ludox solutions were added in Np/Ns ratios ranging from 10-4 to 102, where Ns is the number of silica particles and Np is the number of peptide molecules. The pH of the solutions was checked to be constant during the reactions. QELS measurements were undertaken 15 min, 2 h, and 24 h after mixing and filtration at 500 µm to avoid possible dust interference. 2.4. Charge Titrations. Polyelectrolyte adsorption was monitored after centrifugation of the solution by titration of the remaining polymer with potassium polyvinyl sulfate (KVPS) (Aldrich), using ortho-toluidine blue (o-Tb) as an indicator.13 The charge equivalence of the polycationic peptide was calculated from the amount of KVPS needed to shift the indicator absorption maximum from 635 to 520 nm. 2.5. Solid Characterization. Gels and flocculated solids were collected by centrifugation and dried at room temperature under (11) Allen, L. H.; Matijevic, E. J. Colloid Interface Sci. 1970, 33, 420-429. (12) Mizutani, T.; Nagase, H.; Ogoshi, H. Chem. Lett. 1998, 133134. (13) Horn, D.; Heuck, C. C. J. Biol. Chem. 1983, 258, 1665-1670.
Figure 1. Condensation of silicic acid in the presence of amino acids and peptides. Evolution with time of silicic acid content ([Si] ) 10-2 M, pH ) 7.2) as measured by the optical density ratio OD(t)/OD(t ) 0) at 400 nm using the molybdosilicate method. vacuum. The porosity of the gels or flocculated solids was obtained by the nitrogen sorption experiments performed at 77 K with a Micrometrics 2010 sorptometer. Prior to analysis, samples were first degassed at 60 °C under a 3 µmHg pressure. Specific surface areas (SBET) were determined by the Brunauer-Emmett-Teller (BET) method in the 0.05-0.3 relative pressure range.14 Porous volume (Vp) and average pore size (Rp) were calculated by the Barrett-Joyner-Halenda (BJH) model on the adsorption branch.15 For scanning electron microscopy (SEM), dry powders were coated with gold in a Balzers Union SCD 40 sputter-coater and studied on a Cambridge Stereoscan 120 instrument at an accelerating voltage of 20 kV.
3. Results 3.1. Sodium Silicate Solutions. Silicic acid is obtained upon acidification and dilution of sodium silicate solutions in a Tris-HCl buffer (pH ) 7.2). A clear solution (10-2 M) is obtained. The amount of silicic acid decreases slowly, but no gelation is observed within the first 3 h (Figure 1). This amount seems to decrease slightly faster in the presence of Lys and Arg, but observed variations remain in the error range of the measurements. The corresponding solutions remain optically clear, and gelation is not observed either. On the contrary, solutions become turbid when (p-Lys)35 and (p-Arg)65 are added. The turbidity increases with time, and the amount of Si(OH)4 decreases more rapidly, especially with (p-Arg)65. Moreover, a plateau seems to be reached after 2 h while a white loose gel is formed. Figure 2 shows that the amount of silicic acid decreases more rapidly as the length of the (p-Lys)n chain increases. A white precipitate even forms readily as soon as (pLys)1005, which is poorly soluble in water, is added. The silicic acid content first decreases suddenly, followed by a very slow decay. Nitrogen sorption investigations performed on the dry gels show that they are mesoporous with pore diameters in the 60-70 Å range (Table 1). Comparison with silica gels obtained by adding NaCl (0.1 M) to a silicate solution (10-2 M) in Tris-HCl buffer shows that the presence of polyelectrolytes decreases the surface area but leaves total pore volume constant. Moreover, it appeared that porosity did not depend noticeably on the length of peptide chains. Accordingly, SEM shows that all gels exhibit a similar (14) Brunauer, S.; Emmett, P. H.; Teller, E. J. Am. Chem. Soc. 1938, 60, 309-319. (15) Barrett, E. P.; Joyner, L. G.; Halenda, P. H. J. Am. Chem. Soc. 1951, 73, 373-380.
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Figure 2. Condensation of silicic acid in the presence of polypeptides p-(Lys)n of different chain lengths. Evolution with time of silicic acid content ([Si] ) 10-2 M, pH ) 7.2) as measured by the optical density ratio OD(t)/OD(t ) 0) at 400 nm using the molybdosilicate method. Table 1. BET Surfaces (SBET), Porous Volume (Vp), and Average Pore Size (Rp) As Calculated Using the BJH Model
SBET (m2 g-1) Vp (cm3 g-1) Rp (Å)
silicate/ NaCl
silicate/ p-Lys
Ludox/ NaCl
Ludox/ p-Lys
300 0.45 55
250 0.45 65
150 0.35 85
170 0.70 150
microstructure (Figure 3a). They are formed of agglomerated quasi-spherical particles (diameter < 0.1 µm), as expected from base-catalyzed condensation.16 3.2. Colloidal Silica. Silica sols are added to peptide solutions in R ) Np/Ns ratios. Resulting sols were stable for more than 24 h, until a critical value of R is reached. Beyond this value, flocculation occurs and a white gel is rapidly formed. Similar experiments in the presence of the same amount of the corresponding amino acids show no evolution of the initial silica sol. Figure 4 shows that the average hydrodynamic diameter DH, measured by QELS, does not change significantly until a critical value of R ) Np/Ns ratio is reached. The intersection of the DH ) 16 nm line (corresponding to the initial size of Ludox particles) with the slope of the rapid increase in hydrodynamic diameter preceding flocculation provides an estimate (within a 10-20% error) of a flocculation critical ratio (FCR). FCR is seen to decrease when the polymer chain length increases (Table 2). Analysis of the QELS data (not shown here) suggests that these values correspond to two populations of small (2030 nm) and large (50-500 nm) particles. The relative amount of large particles increases with Np/Ns. After flocculation, QELS of the supernatant shows no signal suggesting that all Ludox particles are flocculated. The peptides remaining in the solution were titrated using KVPS and o-Tb. However, before flocculation, all Ludox particles cannot be removed from the solution, even after centrifugation, and their negative charges interact with the ortho-toluidine blue indicator preventing titration. After flocculation, all Ludox particles are in the solid phase and titration becomes possible showing that no traces of peptides could be detected in the supernatant solution, taking into account the 10 µmol L-1 detection limit of the titration method. Nitrogen adsorption-desorption investigations were performed on the dried flocculates (Table 1). They do not (16) Brinker, C. J.; Scherer, G. W. Sol-Gel Science; Academic Press: San Diego, 1990.
Figure 3. Scanning electron micrographs of (a) silicate/p(Lys)155 xerogel and (b) Ludox/p-(Lys)155 dry flocculated solid.
Figure 4. Evolution of the average hydrodynamic radius DH measured by QELS of a Ludox sol in the presence of polypeptides as a function of the Np/Ns ratio.
depend on the peptide chain length and are characteristic of a meso-to-macroporous silica, as could be expected from large particle flocculation. Comparison with silica precipitated by adding NaCl to Ludox sols shows that in the presence of peptides the specific surface area remains constant whereas total pore volume is increased. Dried flocculated solids are made of large particle aggregates (0.5-1 µm) as shown by SEM (Figure 3b).
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Table 2. Flocculation Critical Ratio of Ludox in the Presence of Different Polyelectrolytes Calculated as Np/Ns and Naa/Nsa p-(Lys)20 p-(Lys)35 p-(Lys)155 p-(Lys)1005 p-(Arg)65 FCR (Np/Ns) FCR (Naa/Ns) a
2 40
0.5 17
0.1 15
0.015 15
0.25 15
Values are given with an estimated 10-20 % error range.
4. Discussion 4.1. Sodium Silicate Solutions. Basic sodium silicate solutions (pH ≈ 11) are known to contain a large number of polysilicate anions such as [Si4O8(OH)4].4-17 Two opposite processes occur when such solutions are diluted and acidified: decondensation occurs upon dilution giving monomeric species [Si(OH)4-x]x-, whereas protonation favors oxolation reactions between Si-OH silanol groups leading to the formation of oligomers and the precipitation of SiO2. Two main species are observed in diluted solutions (10-2 M in Si) around pH ≈ 7, namely, [SiO(OH)3]- and Si(OH)4, silicic acid being the predominant species.18 The condensation kinetics has been shown to increase with pH in the 4-9 pH range.19 Around pH ≈ 7, condensation is supposed to proceed via the reaction of singly ionized species with silicic or polysilicic acid as follows:
Si(OH)4 + [SiO(OH)3]- w (HO)3Si-O-Si(OH)3 + OHIn the absence of amino acids or peptides, the amount of silicic acid decreases slowly with time and gelation is not observed, even after 2 weeks. The presence of NH3+containing amino acids and peptides clearly enhances condensation. A comparison between Lys and Arg with one and three amine side groups, respectively, shows that the total number of amino groups available is an important factor. However, the study of the effect of (p-Lys)n chain length shows that the amount of silicic acid decreases faster when n increases, even though the initial monomerto-Si ratio, n/[Si], is kept constant (Figure 2). The silicic acid decay corresponds to the formation of silicate oligomers.20 Taking the initial slope v of silicic acid decay (see Figure 2) as a measure of the reaction kinetics, ln v can be plotted as a function of m (Figure 5), where m is the number of amino groups per peptide chain (m ) n for p-Lys and m ) 3n for p-Arg).21 Figure 5 shows that ln v increases rapidly at low m values and reaches a plateau for high m values. An interesting parallel could be drawn between these results and published data on the adsorption of charged surfactants onto polyelectrolytes of opposite charge.21 It was shown that the critical aggregation concentration of the surfactants decreases when the polyelectrolyte chain length increases. The Satake and Yang model was then applied, taking into account the binding constant K of the surfactant onto the polyelectrolyte and the cooperative interaction u between the adsorbed surfactant.22 As a matter of fact, K is related to the electric potential at the polyelectrolyte surface. It was shown to depend on the (17) Knight, C. T. G. J. Chem. Soc., Dalton Trans. 1988, 1457-1460. (18) Baes, C. F.; Mesmer, R. E. The hydrolysis of cations; Wiley: New York, 1974. (19) Iler, R. K. The Chemistry of Silica: Solubility, Polymerization, Colloid and Surfaces Properties, and Biochemistry; Wiley: New York, 1979. (20) Harrison, C. C.; Loton, N. J. Chem. Soc., Faraday Trans. 1995, 91, 4287-4297. (21) Liu, J.; Takisawa, N.; Shirahama, K.; Abe, H.; Sakamoto, K. J. Phys. Chem. B 1997, 101, 7520-7523. (22) Satake, I.; Yang, J. T. Biopolymers 1976, 15, 2263-2275.
Figure 5. Variation of ln v as a function of the number m of amino groups per peptide chain in the presence of p-(Lys) (open circle) and p-(Arg) (full circle). v is the initial slope of silicic acid decay.
polymer chain length as long as the number of binding sites is less than a given value, similar to the evolution of the initial slope values reported here.21 In our case, the electrostatic interaction of positively charged amino groups with a negatively charged silica oligomer depends on NH3+-SiO- distance. As a consequence, above a critical chain length, additional positive groups are too far to contribute to the electrostatic potential undergone by the silica anions, thus explaining the plateau illustrated in Figure 5. If such a comparison is valid, it suggests that the observed silicic acid decays can be, at least partially, related to the fixation of silicate anions onto positively charged peptide chains. A cooperative effect could then arise, bringing molecular precursors close enough for condensation to occur all along the peptide chain. Nevertheless, it should be pointed out that the polyelectrolytesilica oligomer interaction is not due to electrostatic interactions only and hydrogen bonds can also be involved. Arginine contains a guanidinium group that carries only one positive charge at pH ) 7, this charge being shared between the three neighboring amino groups. Figure 5 suggests that these three groups are involved in the promotion of silica condensation, so that hydrogen bonding between Si-OH and NH2 groups may also be involved, as suggested earlier.10 This mechanism may not apply to very long chain peptides such as (p-Lys)1005, the global charge of which strongly decreases upon adsorption. Its solubility therefore decreases quickly, leading to precipitation before gelation could occur. SEM pictures of dried solids show that particulate gels are formed. Above pH 7, the polycondensation of silica is known to occur via a monomer-cluster rather than a cluster-cluster aggregation process.16 If the silicapolyelectrolyte model of interactions described above is valid, then one should expect that peptides favor the first steps of the condensation giving oligomers. These oligomers could then behave as nuclei for further condensation via monomer-cluster aggregation until a gel forms. 4.2. Silica Sols. The interaction of charged particles with polyelectrolytes of opposite charge is already well documented.23-25 The effect of particle size26 and polymer molecular weight27 was studied in model28-30 and biological (23) Healy, T. W.; La Mer, V. K. J. Colloid Sci. 1964, 19, 323-332. (24) Heller, W. Pure Appl. Chem. 1966, 12, 249-274. (25) Vincent, B. Adv. Colloid Interface Sci. 1974, 4, 193-277. (26) Iler, R. K. J. Colloid Interface Sci. 1971, 37, 364-373. (27) Lindquist, G. M.; Stratton, R. A. J. Colloid Interface Sci. 1976, 55, 45-59. (28) Walker, H. W.; Grant, S. B. Colloids Surf., A 1996, 119, 229239. (29) Poncet-Legrand, C.; Lafuma, F.; Audebert, R. Colloids Surf., A 1999, 152, 251-261.
Interactions of Peptides with Silica
Langmuir, Vol. 18, No. 6, 2002 2335 Scheme 1. Schematic Illustration of Proposed Models for (a) Silica Gelation in the Silicate/p-Lys System and (b) Silica Flocculation in the Ludox/p-Lys System
Figure 6. Effect of peptide chain length n on the flocculation critical ratio FCR of Ludox in the presence of p-(Lys) (open circle) and p-(Arg) (full circle).
systems.31-33 However, to the best of our knowledge, specific studies of p-Lys and p-Arg interactions with diluted silica sols and comparison with silicate solutions have never been reported. At small Np/Ns ratios, QELS observations suggest the coexistence of two populations: isolated Ludox particles and aggregates made of silica particles bound to a polypeptide chain via Si-O-/NH3+ electrostatic interactions. In each aggregate, the polypeptide chain carries a constant number of silica particles. The number of aggregates increases with the number of peptide chains, until a critical ratio, FCR, is reached when there are enough peptides to bind all silica particles. If charges are still available on the particle surface, extra polypeptides will also tend to bind, through electrostatic interactions, as many particles as possible, giving rise to the possibility of bridge formation between intra-aggregate particles.26 This leads to destabilization of the sols and flocculation. The efficiency of this process will partly depend on polymer possible spatial extension, which is correlated to polymer chain length.27 The fact that the supernatant solution after flocculation contains neither silica particles nor polymers reinforces this model. Moreover, analysis of dry flocculated solids, indicating porous volume and pore size about 2 times larger than in the presence of NaCl but with a similar specific surface area, is in agreement with the existence of larger aggregates associated in a loose network.19 FCR values can also give an indication of the number of silica particles bound to each polypeptide chain within an aggregate. FCR is merely inversely proportional to the polyelectrolyte degree of condensation, as illustrated in Figure 6, except for shortest chain (n ) 20). As a consequence, when FCR is recalculated as the Naa/Ns ratio, where Naa is the number of amino acids (Naa ) Npn) (Table 2), this ratio does not seem to depend on the polymer size. Moreover, it allows us to conclude that only one of the three available amino groups of p-Arg is able to bind silica particles. The corresponding FCR value of 15 amino acids per particle is very close to the p-(Lys)20 chain length. This would imply that each p-(Lys)20 can only bind one particle. However, in this case, FCR values indicate an average of 2 polymer chains (or 35-40 amino acids) per (30) Schwarz, S.; Lunkwitz, K.; Kessler, B.; Spiegler, U.; Killmann, E.; Jaeger, W. Colloids Surf., A 2000, 163, 17-27. As a matter of fact, decreasing the pH of the silica sols from 7 to 5 in the absence of polymer does not lead to flocculation, even though the surface charge is lowered to about 80 negative charges per particle. (31) Haynes, C.; Sliwinsky, E.; Norde, W. J. Colloid Interface Sci. 1994, 164, 394-409. (32) Karlsson, M.; Martensson, L. G.; Jonsson, B. H.; Carlsson, U. Langmuir 2000, 16, 8470-8479. (33) Hiddessen, A. L.; Rodgers, S. D.; Weitz, D. A.; Hammer, D. A. Langmuir 2000, 16, 9744-9753.
particle. This suggests that once every polymer is bound to one Ludox particle, an additional chain is not long enough to form a bridge by sharing its binding sites between two particles but remains located at the surface of a single particle. Finally, considering that a maximum of ca. 15 positive NH3+ groups can interact with the 220240 negative charges of the Ludox particles, this clearly indicates that the polyelectrolyte-silica aggregates are still highly charged so that precipitation due to decrease of their global ζ potential can be excluded.30 On the contrary, it suggests that enough negatively charged surface is available for further bridging of aggregates one with another by additional polymer. 5. Conclusion This work studies the effect of amino-containing polypeptides on silicic acid solutions and silica sols. In both cases, it is suggested that electrostatic interactions between positively charged amino groups of the peptide chain and negatively charged silica species lead to rapid precipitation. The major role of polymer chain length underlines that the number of NH3+ binding sites is a key parameter, suggesting that polypeptides may serve as substrates for silica formation. However, two different processes should occur depending on the nature of silicate species. In the case of sodium silicate solution, silicic acid monomers are brought close enough by interaction with amino groups to favor oligomerization. Gelation then proceeds by further condensation of monomers with these precondensed species (Scheme 1a). In contrast, silica particles are bound to the polyelectrolyte chains to form peptide-silica assemblies in which strong interparticle interactions are not expected. Silica precipitation then occurs via the bridging of aggregates by additional polymer (Scheme 1b). Coming back to biogenic silica, silaffins have been described as low molecular weight proteins containing Lys-Lys and Arg-Arg clusters regularly distributed along the peptide backbone.7 Moreover, they have been shown
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to precipitate silicic acid solution to form silica spheres with diameters of ca. 500 nm or aggregates of smaller particles (diameters < 50 nm). As suggested from porosity measurements and SEM studies, silicate condensation leads to mesoporous solids made via the aggregation of small particles (
