The structure-function relationship of the lipases from Pseudomonas aeruginosa and Bacillus subtilis

Onno Misset, Gijs Gerritse, Karl Erich Jaeger, Ulrich Winkler, Charles Colson, Karin Schanck, Emmanuel Lesuisse, Véronique Dartois, Mieke Blaauw, Stéphane Ransac, Bauke W. Dijkstra

Research output: Contribution to journalArticlepeer-review

33 Scopus citations

Abstract

Within the BRIDGE T-project on lipases we investigate the structure-function relationships of the lipases from Bacillus subtilis and Pseudomonas aeruginosa. Construction of an overproducing Bacillus. strain allowed the purification of > 100 mg lipase from 30 l culture supernatant. After testing a large variety of crystallization conditions, the Bacillus lipase gave crystals of reasonable quality in PEG-4000 (38-45%), Na2SO4 and octyl-β-glucoside at 22°C, pH 9.0. A 2.5 Å; dataset has been obtained which is complete from 15 to 2.5 A resolution. P.aeruginosa wild-type strain PAC1R was fermented using conditions of maximum lipase production. More than 90% of the lipase was cell bound and could be solubilized by treatment of the cells with Triton X-100. This permitted the purification of ∼50 mg lipase. So far, no crystals of sufficient quality were obtained. Comparison of the model we built for the Pseudomonas lipase, on the basis of sequences and structures of various hydrolases which were found to possess a common folding pattern (α/β hydrolase fold), with the X-ray structure of the P.glumae lipase revealed that it is possible to correctly build the structure of the core of a protein even in the absence of obvious sequence homology with a protein of known 3-D structure.

Original languageEnglish (US)
Pages (from-to)523-529
Number of pages7
JournalProtein Engineering, Design and Selection
Volume7
Issue number4
DOIs
StatePublished - Apr 1994
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • General Medicine

Keywords

  • 3-D structure
  • Bacillus subtilis
  • Crystallization
  • Lipase
  • Pseudomonas aeruginosa

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