Title
Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop
Document Type
Article
Publication Date
11-1-2007
Abstract
Lipoxygenases (LOXs) catalyze the regiospecific and stereospecific dioxygenation of polyunsaturated membrane-embedded fatty acids. A Ca(2+)-dependent membrane-binding function was localized to the amino-terminal C2-like domain of 8R-lipoxygenase (8R-LOX) from the soft coral Plexaura homomalla. The 3.2 A crystal structure of 8R-LOX and spectroscopic data suggested that Ca(2+) stabilizes two membrane-insertion loops. Analysis of the protein packing contacts in the crystal lattice indicated that the conformation of one of the two loops complicated efforts to improve the resolution of the X-ray data. A deletion mutant of 8R-LOX in which the corresponding membrane-insertion loop is absent (Delta41-45:GSLOX) was engineered. Removal of the membrane-insertion loop dramatically increases the protein yield from bacterial cultures and the quality of the crystals obtained, resulting in a better than 1 A improvement in the resolution of the diffraction data.
Publication Source (Journal or Book title)
Acta crystallographica. Section F, Structural biology and crystallization communications
First Page
972
Last Page
5
Recommended Citation
Neau, D. B., Gilbert, N. C., Bartlett, S. G., Dassey, A., & Newcomer, M. E. (2007). Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop. Acta crystallographica. Section F, Structural biology and crystallization communications, 63 (Pt 11), 972-5. https://doi.org/10.1107/S1744309107050993