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Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop.

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.

SUBMITTER: Neau DB

PROVIDER: S-EPMC2339740 | biostudies-literature | 2007 Nov

REPOSITORIES: biostudies-literature

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Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop.

Neau David B DB Gilbert Nathaniel C NC Bartlett Sue G SG Dassey Adam A Newcomer Marcia E ME

Acta crystallographica. Section F, Structural biology and crystallization communications 20071024 Pt 11

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 th ...[more]

PMID: 18007054

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Dataset Information

Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop.

Publications

Improving protein crystal quality by selective removal of a Ca(2+)-dependent membrane-insertion loop.

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