Over-Production, Crystallization, and Preliminary X-ray Crystallographic Analysis of a Coiled-Coil Region in Human Pericentrin

초록

The genes encoding three coiled-coil regions in human pericentrin were gene synthesized with Escherichia coli codon-optimization, and the proteins were successfully over-produced in large quantities using E. coli expression. After verifying that the purified proteins were mostly composed of -helices, one of the proteins was crystallized using polyethylene glycol 8000 as crystallizing agent. X-ray diffraction data were collected to 3.8 angstrom resolution under cryo-condition using synchrotron X-ray. The crystal belonged to space group C2 with unit cell parameters a = 324.9 angstrom, b = 35.7 angstrom, c = 79.5 angstrom, and = 101.6. According to Matthews' coefficient, the asymmetric unit may contain up to 12 subunits of the monomeric protein, with a crystal volume per protein mass (V-M) of 1.96 angstrom(3) Da(-1) and a 37.3% solvent content.

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