In‐situ 2D bacterial crystal growth as a function of protein concentration: An atomic force microscopy study

• Published in: Microscopy research and technique Vol. 81; no. 10; pp. 1095 - 1104
• Main Authors: Moreno‐Cencerrado, Alberto, Iturri, Jagoba, Toca‐Herrera, José L.
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01.10.2018; John Wiley and Sons Inc
• Subjects: Atomic force microscopy; Avrami equation; Bacillaceae - metabolism; Bacterial Proteins - chemistry; Crystal growth; Crystal lattices; Crystallization; Crystals; Elastic Modulus - physiology; Fluorides; Growth rate; high‐resolution atomic force microscopy; Hydrophobicity; Lattice parameters; Microscopy; Microscopy, Atomic Force - methods; Monosaccharide Transport Proteins - chemistry; Nucleation; protein adsorption; Protein crystal growth; Protein interaction; Proteins; protein‐substrate interaction; Silicon dioxide; Substrates; Unit cell; Viscoelasticity; protein adsorption; high-resolution atomic force microscopy; crystal growth; protein-substrate interaction; Avrami equation
• ISSN: 1059-910X; 1097-0029
• DOI: 10.1002/jemt.23075

The interplay between protein concentration and (observation) time has been investigated for the adsorption and crystal growth of the bacterial SbpA proteins on hydrophobic fluoride‐functionalized SiO2 surfaces. For this purpose, atomic force microscopy (AFM) has been performed in real‐time for monitoring protein crystal growth at different protein concentrations. Results reveal that (1) crystal formation occurs at concentrations above 0.08 µM and (2) the compliance of the formed crystal decreases by increasing protein concentration. All the crystal domains observed presented similar lattice parameters (being the mean value for the unit cell: a = 14.8 ± 0.5 nm, b = 14.7 ± 0.5 nm, γ = 90 ° ± 2). Protein film formation is shown to take place from initial nucleation points which originate a gradual and fast extension of the crystalline domains. The Avrami equation describes well the experimental results. Overall, the results suggest that protein‐substrate interactions prevail over protein–protein interactions. Research Highlights AFM enables to monitor protein crystallization in real‐time. AFM high‐resolution determines lattice parameters and viscoelastic properties. S‐layer crystal growth rate increases with protein concentration. Avrami equation models protein crystal growth. Figure Legend: 2D protein crystal growth. On the upper row, the crystalline protein domains grow in an isotropic manner along the surface. Below, the protein area (in blue) is simulated by means of digital imaging process.