The isopenicillin-N Acyltransferase of Penicillium Chrysogenum Has isopenicillin-N Amidohydrolase, 6-aminopenicillanic Acid Acyltransferase and Penicillin Amidase Activities, All of Which Are Encoded by the Single penDE Gene

Eur J Biochem. 1993 Jul 15;215(2):323-32. doi: 10.1111/j.1432-1033.1993.tb18038.x.

Abstract

The isopenicillin-N acyltransferase of Penicillium chrysogenum catalyzes the conversion of the biosynthetic intermediate isopenicillin N to the hydrophobic penicillins. The isopenicillin-N acyltransferase copurified with the acyl-CoA:6-aminopenicillanic acid (6-APA) acyltransferase activity which transfers an acyl residue from acyl-CoA derivatives (e.g. phenylacetyl-CoA, phenoxyacetyl-CoA) to 6-APA. Other thioesters of phenylacetic acid were also used as substrates. An amino acid sequence similar to that of the active site of thioesterases was found in the isopenicillin-N acyltransferase, suggesting that this site is involved in the transfer of phenylacetyl residues from phenylacetyl thioesters. Purified isopenicillin-N acyltransferase also showed isopenicillin-N amidohydrolase, penicillin transacylase and penicillin amidase activities. The isopenicillin-N amidohydrolase (releasing 6-APA) showed a much lower specific activity than the isopenicillin-N acyltransferase of the same enzyme preparation, suggesting that in the isopenicillin-N acyltransferase reaction the 6-APA is not released and is directly converted into benzylpenicillin. Penicillin transacylase exchanged side chains between two hydrophobic penicillin molecules; or between one penicillin molecule and 6-APA. The penicillin amidase activity is probably the reverse of the biosynthetic acyl-CoA:6-APA acyltransferase. Four P. chrysogenum mutants deficient in acyl-CoA:6-APA acyltransferase lacked the other four related activities. Transformation of these mutants with the penDE gene restored all five enzyme activities.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acyltransferases / genetics
  • Acyltransferases / isolation & purification
  • Acyltransferases / metabolism*
  • Amidohydrolases / isolation & purification
  • Amidohydrolases / metabolism*
  • Amino Acid Sequence
  • Culture Media
  • Dithiothreitol / pharmacology
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme Induction / drug effects
  • Gene Expression
  • Molecular Sequence Data
  • Mutation
  • Penicillin Amidase / genetics
  • Penicillin Amidase / isolation & purification
  • Penicillin Amidase / metabolism*
  • Penicillin-Binding Proteins*
  • Penicillins / metabolism*
  • Penicillium chrysogenum / enzymology*
  • Penicillium chrysogenum / genetics
  • Substrate Specificity
  • Sulfhydryl Reagents / pharmacology

Substances

  • Culture Media
  • Penicillin-Binding Proteins
  • Penicillins
  • Sulfhydryl Reagents
  • Acyltransferases
  • acyl-CoA-6-aminopenicillanic acid acyltransferase
  • Amidohydrolases
  • Penicillin Amidase
  • penicillin N
  • Dithiothreitol