Molecular mechanisms of microheterogeneity-induced defect formation in ferritin crystallization.
We apply in situ atomic force microscopy to the crystallization of ferritins from solutions containing approximately 5% (w/w) of their inherent molecular dimers. Molecular resolution imaging shows that the dimers consist of two bound monomers. The constituent monomers are likely partially denatured, resulting in increased hydrophobicity of the dimer surface. Correspondingly, the dimers strongly adsorb on the crystal surface. The adsorbed dimers hinder step growth and on incorporation by the crystal initiate stacks of up to 10 triple and single vacancies in the subsequent crystal layers. The molecules around the vacancies are shifted by approximately 0.1 molecular dimensions from their crystallographic positions. The shifts strain the lattice and, as a consequence, at crystal sizes > 200 microm, the accumulated strain is resolved by a plastic deformation whereupon the crystal breaks into mosaic blocks 20-50 microm in size. The critical size for the onset of mosaicity is similar for ferritin and apoferritin and close to the value for a third protein, lysozyme; it also agrees with theoretical predictions. Trapped microcrystals in ferritin and apoferritin induce strain with a characteristic length scale equal to that of a single point defect, and, as a consequence, trapping does not contribute to the mosaicity. The sequence of undesired phenomena that include heterogeneity generation, adsorption, incorporation, and the resulting lattice strain and mosaicity in this and other proteins systems, could be avoided by improved methods to separate similar proteins species (microheterogeneity) or by increasing the biochemical stability of the macromolecules against oligomerization.
Yau
S T
ST
Center for Microgravity and Materials Research, University of Alabama in Huntsville, Huntsville, Alabama 35899, USA.
Thomas
B R
BR
Galkin
O
O
Gliko
O
O
Vekilov
P G
PG
D000327:Adsorption; D001052:Apoferritins / Q000737:chemistry*; D055592:Biophysical Phenomena; D001703:Biophysics; D003460:Crystallization; D019281:Dimerization; D005293:Ferritins / Q000737:chemistry*; D066298:In Vitro Techniques; D046911:Macromolecular Substances; D018625:Microscopy, Atomic Force; D011487:Protein Conformation; D013816:Thermodynamics
D046911:Macromolecular Substances; D005293:Ferritins; D001052:Apoferritins