Species-specific polyamines from diatoms control silica morphology

Abstract

Biomineralizing organisms use organic molecules to generate species-specific mineral patterns. Here, we describe the chemical structure of long-chain polyamines (up to 20 repeated units), which represent the main organic constituent of diatom biosilica. These substances are the longest polyamine chains found in nature and induce rapid silica precipitation from a silicic acid solution. Each diatom is equipped with a species-specific set of polyamines and silica-precipitating proteins, which are termed silaffins. Different morphologies of precipitating silica can be generated by polyamines of different chain lengths as well as by a synergistic action of long-chain polyamines and silaffins.

Figure 1

HF extracts from diatom cell walls. Extracts were subjected to Tris-Tricine SDS/PAGE (11) and stained with Coomassie blue. Lanes: 1, C. fusiformis (positions of silaffin species are marked); 2, N. angularis; 3, C. didymum; 4, C. debilis; 5, E. zodiacus; 6, S. turris. The brackets in lane 2 mark the three silaffins that were used for silica precipitation (see Fig. 6).

Figure 2

Characterization of diatom polyamines. Electrospray ionization/MS analysis of purified polyamine fractions. Each peak represents a singly charged positive ion. Selected peaks are marked by their m/z units. (A) C. fusiformis polyamines. (B) C. didymum polyamines. (C) N. angularis polyamines.

Figure 3

Permethylation analysis of the polyamines from N. angularis. (A and B) Electrospray ionization/MS analysis of singly charged positive ions. (A) Unmodified polyamines from N. angularis. Two peak series, which are separated by 71 units, are denoted by vertical numbers that indicate the m/z value of the corresponding peak. Within each series, peak masses differ by 14 units. The horizontal numbers indicate selected molecules whose masses differ by 71 units. (B) Polyamines from N. angularis after permethylation. Two peak series can be discerned, which are denoted by horizontal numbers (putrescine basis) and vertical numbers (ornithine basis), respectively. Within each series, peak masses differ by 71 units. (C) Scheme of proposed general polyamine structure. The gray box highlights the putrescine moiety. (D) Theoretical molecular masses of putrescine-based polyamine molecules. Each line corresponds to a given polyamine chain length and lists the theoretical molecular masses of methylation isoforms. The molecular masses of fully methylated isoforms are boxed.

Figure 4

Fragmentation analysis. (A) Product ion spectrum obtained by collision-induced fragmentation of the (m + H)⁺ = 856 ion (see mass signal in Fig. 3B). Two series of ions were detected that differ by 14 units. Within each series, neighboring peaks differ by 71 units. (B) Proposed structure (schematic) of the (m + H)⁺ = 856 ion. Cleavage positions that lead to the observed fragment ions are depicted by rectangular arrows, and the corresponding masses are indicated. The putrescine residue is highlighted by a gray box.

Figure 5

Silica precipitates induced by N. angularis polyamines. For precipitation, polyamines of molecular masses from 1,000 to 1,250 (A) and 600 to 750 Da (B) were used. (C–F) The natural mixture of polyamines (molecular masses 600–1,250 Da) was used for investigating the pH dependence on polyamine-induced silica precipitation. (C) pH 5.4. (D) pH 6.3. (E) pH 7.2. (F) pH 8.3. The polyamine concentration in each solution was 0.85 mg/ml. [Scale bars, 1 μm (A and B) and 500 nm (C–F).]

Figure 6

Effect on silica morphology of combining N. angularis polyamines and silaffins. For precipitation in A, enriched silaffins (see brackets in Fig. 1, lane 2) at a concentration of 3 mg/ml were used and in B, a mixture of silaffins (3 mg/ml) and polyamines (0.85 mg/ml) was used. (Insets) Larger views of the precipitates. [Scale bars, 500 nm, 1 μm (Insets).]