Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis
Journal Article
·
· Acta Crystallographica. Section F
- Division of Biochemical Sciences, National Chemical Laboratory, Pune 411 008 (India)
- Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5YW (United Kingdom)
An unannotated protein reported from B. subtilis has been expressed in E. coli and identified as possessing penicillin V acylase activity. The crystallization and preliminary crystallographic analysis of this penicillin V acylase is presented. Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β-lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging-drop vapour-diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C222{sub 1}, with unit-cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å{sup 3} Da{sup −1}, corresponding to 62% solvent content. The structure has been solved using molecular-replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model.
- OSTI ID:
- 22356140
- Journal Information:
- Acta Crystallographica. Section F, Journal Name: Acta Crystallographica. Section F Journal Issue: Pt 7 Vol. 61; ISSN ACSFCL; ISSN 1744-3091
- Country of Publication:
- United Kingdom
- Language:
- English
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