title: Purification, crystallization and preliminary X-ray diffraction of Cys103Ala acyl coenzyme A: isopenicillin N acyltransferase from Penicillium chrysogenum.
abstract: Penicillins and cephalosporins are an efficacious group of antibiotics produced by fungi such as Penicillium chrysogenum and Acremonium chrysogenum. The last step in their biosynthesis is catalyzed by acyl coenzyme A:isopenicillin N transferase (AT). This enzyme is produced as a single-chain proenzyme, which is activated by autocatalytic cleavage of the Gly102-Cys103 peptide bond, resulting in a heterodimeric protein with subunits of 11 and 29 kDa. The Cys103Ala mutant of the proenzyme, which does not undergo this cleavage, was purified and crystallized. Diffraction-quality crystals of the mutant and an L-SeMet-substituted mutant were obtained by vapour diffusion against solutions containing (NH(4))(2)SO(4), NaCl and HEPES-NaOH pH 7.5. The crystals belong to the monoclinic space group C2, with unit-cell parameters a = 231.36, b = 68.27, c = 151.31 A and beta = 129.56 degrees. They diffract to 2.8 A resolution with X-rays from a rotating-anode generator.
authors.affiliation: University of Groningen, Laboratory of Biophysical Chemistry, Nijenborgh 4, 9747 AG Groningen, The Netherlands.
authors.forename: Charles M H
authors.initials: CM
authors.lastname: Hensgens
authors.forename: Els A
authors.initials: EA
authors.lastname: Kroezinga
authors.forename: Bart A
authors.initials: BA
authors.lastname: van Montfort
authors.forename: Jan-Metske
authors.initials: JM
authors.lastname: van der Laan
authors.forename: John D
authors.initials: JD
authors.lastname: Sutherland
authors.forename: Bauke W
authors.initials: BW
authors.lastname: Dijkstra
mesh_terms: D000217:Acyltransferases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D019943:Amino Acid Substitution; D003460:Crystallization; D013058:Mass Spectrometry; D046915:Penicillin-Binding Proteins; D010408:Penicillium chrysogenum / Q000737:chemistry*; D014961:X-Ray Diffraction
chemical_list: D046915:Penicillin-Binding Proteins; D000217:Acyltransferases; C018756:acyl-CoA-6-aminopenicillanic acid acyltransferase