A common feature of biomineralization proteins is their self-assembly to produce a surface consistent in size with the inorganic crystals that they produce. Mms6, a small protein of 60 amino acids from Magnetospirillum magneticum strain AMB-1 that promotes the in vitro growth of superparamagnetic magnetite nanocrystals, assembles in aqueous solution to form spherical micelles that could be visualized by TEM and AFM. The results reported here are consistent with the view that the N and C-terminal domains interact with each other within one polypeptide chain and across protein units in the assembly. From studies to determine the amino acid residues important for self-assembly, we identified the unique GL repeat in the N-terminal domain with additional contributions from amino acids in other positions, throughout the molecule. Analysis by CD spectroscopy identified a structural change in the iron-binding C-terminal domain in the presence of Fe3+. A change in the intrinsic fluorescence of tryptophan in the N-terminal domain showed that this structural change is transmitted through the protein. Thus, self-assembly of Mms6 involves an interlaced structure of intra- and inter-molecular interactions that results in a coordinated structural change in the protein assembly with iron binding.

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Hydrophobins are a group of low-molecular-mass, cysteine-rich proteins that have unusual biophysical properties. They are highly surface-active and can self-assemble at hydrophobic-hydrophilic interfaces, forming surface layers that are able to reverse the hydropathy of surfaces. Here we describe a novel hydrophobin from the edible mushroom Grifola frondosa, which was named HGFI and belongs to class I. The hydrophobin gene was identified during sequencing of random clones from a cDNA library, and the corresponding protein was isolated as a hot SDS-insoluble aggregate from the cell wall. The purified HGFI was found to have 83 amino acids. The protein sequence deduced from the cDNA sequence had 107 amino acids, from which a 24 aa signal sequence had been cleaved off in the mature protein. This signal sequence was 5 aa longer than had been predicted on the basis of signal peptide analysis of the cDNA. Rodlet mosaic structures were imaged using atomic force microscopy (AFM) on mica surfaces after drying-down HGFI solutions. Using Langmuir films we were also able to take images of both the hydrophobic and hydrophilic sides of films formed at the air-water interface. No distinct structure was observed in films compressed once, but in films compressed several times rodlet structures could be seen. Most rodlets were aligned in the same direction, indicating that formation of rodlets may be promoted during compression of the monolayer.

Magnetotactic bacteria that produce magnetic nanocrystals of uniform size and well-defined morphologies have inspired the use of biomineralization protein Mms6 to promote formation of uniform magnetic nanocrystals in vitro. Small angle X-ray scattering (SAXS) studies in physiological solutions reveal that Mms6 forms compact globular three-dimensional (3D) micelles (approximately 10 nm in diameter) that are, to a large extent, independent of concentration. In the presence of iron ions in the solutions, the general micellar morphology is preserved, however, with associations among micelles that are induced by iron ions. Compared with Mms6, the m2Mms6 mutant (with the sequence of hydroxyl/carboxyl containing residues in the Cterminal domain shuffled) exhibits subtle morphological changes in the presence of iron ions in solutions. The analysis of the SAXS data is consistent with a hierarchical core-corona micellar structure similar to that found in amphiphilic polymers. The addition of ferric and ferrous iron ions to the protein solution induces morphological changes in the micellar structure by transforming the 3D micelles into objects of reduced dimensionality of 2, with fractal-like characteristics (including Gaussian-chain-like) or, alternatively, plateletlike structures. Small angle X-ray scattering (SAXS) studies in physiological solutions reveal that Mms6 forms compact globular threedimensional (3D) micelles (approximately 10 nm in diameter) that are, to a large extent, independent of concentration. In the presence of iron ions in the solutions, the general micellar morphology is preserved, however, with associations among micelles that are induced by iron ions. Compared with Mms6, the m2Mms6 mutant (with the sequence of hydroxyl/carboxyl containing residues in the C-terminal domain shuffled) exhibits subtle morphological changes in the presence of iron ions in solutions. The analysis of the SAXS data is consistent with a hierarchical core−corona micellar structure similar to that found in amphiphilic polymers. The addition of ferric and ferrous iron ions to the protein solution induces morphological changes in the micellar structure by transforming the 3D micelles into objects of reduced dimensionality of 2, with fractal-like characteristics (including Gaussian-chain-like) or, alternatively, platelet-like structures.