Interfacial Properties and Iron Binding to Bacterial Proteins That Promote the Growth of Magnetite Nanocrystals: X-ray Reflectivity and Surface Spectroscopy Studies

Abstract

Surface sensitive X-ray scattering and spectroscopic studies have been conducted to determine structural properties of Mms6, the protein in Magnetospirillum magneticum AMB-1 that is implicated as promoter of magnetite nanocrystals growth. Surface pressure versus molecular area isotherms indicate Mms6 forms stable monolayers at the aqueous/vapor interface that are strongly affected by ionic conditions of the subphase. Analysis of X-ray reflectivity from the monolayers shows that the protein conformation at the interface depends on surface pressure and on the presence of ions in the solutions, in particular of iron ions and its complexes. X-ray fluorescence at grazing angles of incidence from the same monolayers allows quantitative determination of surface bound ions to the protein showing that ferric iron binds to Mms6 at higher densities compared to other ions such as Fe 2+ or La 3+ under similar buffer conditions. ABSTRACT: Surface sensitive X-ray scattering and spectroscopic studies have been conducted to determine structural properties of Mms6, the protein in Magnetospirillum magneticum AMB-1 that is implicated as promoter of magnetite nanocrystals growth. Surface pressure versus molecular area isotherms indicate Mms6 forms stable monolayers at the aqueous/vapor interface that are strongly affected by ionic conditions of the subphase. Analysis of X-ray reflectivity from the monolayers shows that the protein conformation at the interface depends on surface pressure and on the presence of ions in the solutions, in particular of iron ions and its complexes. X-ray fluorescence at grazing angles of incidence from the same monolayers allows quantitative determination of surface bound ions to the protein showing that ferric iron binds to Mms6 at higher densities compared to other ions such as Fe 2+ or La 3+ under similar buffer conditions. Disciplines Biochemical and Biomolecular

Genetic Dissection of the mamAB and mms6 Operons Reveals a Gene Set Essential for Magnetosome Biogenesis in Magnetospirillum gryphiswaldense

“…MamN exhibits similarity to H ϩ translocation proteins and might be involved in crystal growth by regulating intramagnetosomal pH (51). Mms6 is tightly bound to the magnetosome crystals (26,28) and assembles into coherent micelles for templating crystal growth (57). mms48 (Mms48) and mms36 (Mms36) act as inhibitors of crystal growth or recruit inhibiting proteins of particle growth by an unknown mechanism.…”

Morphological Transformations in the Magnetite Biomineralizing Protein Mms6 in Iron Solutions: A Small-Angle X-ray Scattering Study

“…19,21,27,28 The C-terminal domains bind ferric iron ions and complexes with very high affinity. 21,28 Shuffling the amino acid sequence in a protein affects its function and thus is a good test to understand the mechanism of biomineralization. A C-terminal mutant, m2Mms6, has been designed that shows much lower iron binding than the wild-type Mms6.…”

“…16,21 The Mms6−iron interactions at various pH values have been previously investigated on aqueous surfaces, where Mms6 was deposited on the surface of an iron solute subphase to form a monolayer, taking advantage of its amphiphilic behavior. 28 In this SAXS study, we investigate the interactions of Mms6 with iron salts in Tris/KCl solutions under very similar conditions to room temperature in vitro synthesis of magnetite. 20,21 The role of pH in the morphology of the protein is also reported as a control in that the presence of iron in solutions affects the pH.…”

Amorphous iron-(hydr) oxide networks at liquid/vapor interfaces: In situ X-ray scattering and spectroscopy studies

“…Recently, a trans-membrane protein called Mms6 has been identified as the promoter of magnetite single crystal growth both in vivo and in vitro by providing acidic and basic regions that are necessary to distinguish Fe (III) from Fe (II) ions [7][8][9][10]. Significant iron-binding to the Mms6 has been observed in Mms6 as a micelle in bulk solutions [11] or as a film on aqueous surfaces [12].…”

“…In this study we used dihexadecyl-phosphate (DHDP) and arachidic acid (AA) to form two-dimensional (2D) charged templates. Our main objective in this study was not to grow crystals at the charged interface, but to determine the iron (Fe (III) and Fe (II)) accumulation near charged surfaces consisting of either phosphate groups or carboxyl groups, as a direct comparison with the iron binding to Mms6 at air/aqueous solution interfaces [12] under otherwise identical subphase conditions. In order to maintain conditions as close as possible to physiological ones that preserve the functionality of Mms6, the DHDP and AA were each spread on subphase solutions consisting of tris (hydroxymethyl) aminomethane (Tris) and KCl (referred to as Tris-KCl) at pH 7.0, to which we added the iron salts (FeCl 2 or FeCl 3 ) and added hydrochloric acid (HCl) to lower the pH to $3, a protocol that is commonly used to promote the iron-oxide crystal in vitro by Mms6 [11,12].…”