Extracellular overproduction and preliminary crystallographic analysis of a family I.3 lipase

Abstract

A family I.3 lipase from Pseudomonas sp. MIS38 was secreted from Escherichia coli cells to the external medium, purified and crystallized and preliminary crystallographic studies were performed. The crystal was grown at 277 K by the hanging-drop vapour-diffusion method. Native X-ray diffraction data were collected to 1.7 A resolution using synchrotron radiation at station BL38B1, SPring-8. The crystal belongs to space group P2(1), with unit-cell parameters a = 48.79, b = 84.06, c = 87.04 A. Assuming the presence of one molecule per asymmetric unit, the Matthews coefficient V(M) was calculated to be 2.73 A3 Da(-1) and the solvent content was 55%.

Figure 1

Crystal of PML grown by the hanging-drop vapour-diffusion method. The approximate dimensions of the crystal are 0.3 × 0.2 × 0.05 mm.

Figure 2

X-ray diffraction pattern of the native PML crystal. The crystal was exposed at 100 K after soaking in artificial mother liquor containing 20% ethylene glycol for cryoprotection. The photograph shows a single frame of 1.0° oscillation with an exposure time of 10 s per image and a crystal-to-detector distance of 170 mm. The crystal diffracted to 1.7 Å (the edge of the plate).