Multistep Crystallization of Microbial S-Layers
Many microbes assemble a crystalline protein layer on their outer surface as an additional barrier and communication platform between the cell and its environment. These surface layer (S-layer) proteins efficiently crystallize to continuously coat the cell, and this trait has been utilized to design functional macromolecular nanomaterials.
A collaborative research team from Stanford University and the University of British Columbia, led by S. Wakatsuki, examined the structural basis for self-assembly of RsaA, a 98-kDa S-layer protein from Caulobacter crescentus, in vitro using a combination of time-resolved small angle x-ray scattering performed at SSRL’s beam line 4-2, circular dichroism spectroscopy, X-ray macromolecular crystallography, and a time course of cryogenic electron microscopy (Cryo-EM).
The team established that the assembly pathway involves two domains serving distinct functions. The C-terminal crystallization domain forms the physiological 2-dimensional (2D) crystal lattice, but full-length protein crystallizes multiple orders of magnitude faster due to the N-terminal nucleation domain.
Crystallization observations using a time course of cryo-EM imaging revealed a crystalline intermediate wherein N-terminal nucleation domains exhibit motional dynamics with respect to rigid lattice-forming crystallization domains. Dynamic flexibility between the two domains rationalizes efficient S-layer crystal nucleation on the curved cellular surface.
The research demonstrates that a discrete nucleation domain is responsible for enhancing the rate of self-assembly, unveiling possible mechanisms to engineer kinetically controllable self-assembling 2D macromolecular nanomaterials.
Herrmann, J., et al. 2020. “A Bacterial Surface Layer Protein Exploits Multistep Crystallization for Rapid Self-Assembly,” Proceedings of the National Academy of Sciences USA 117(1), 388–94. [DOI:10.1073/pnas.1909798116]