Engineering of Penicillium chrysogenum for fermentative production of a novel carbamoylated cephem antibiotic precursor.
Penicillium chrysogenum was successfully engineered to produce a novel carbamoylated cephalosporin that can be used as a synthon for semi-synthetic cephalosporins. To this end, genes for Acremonium chrysogenum expandase/hydroxylase and Streptomyces clavuligerus carbamoyltransferase were expressed in a penicillinG high-producing strain of P.chrysogenum. Growth of the engineered strain in the presence of adipic acid resulted in production of adipoyl-7-amino-3-carbamoyloxymethyl-3-cephem-4-carboxylic acid (ad7-ACCCA) and of several adipoylated pathway intermediates. A combinatorial chemostat-based transcriptome study, in which the ad7-ACCCA-producing strain and a strain lacking key genes in beta-lactam synthesis were grown in the presence and absence of adipic acid, enabled the dissection of transcriptional responses to adipic acid per se and to ad7-ACCCA production. Transcriptome analysis revealed that adipate catabolism in P.chrysogenum occurs via beta-oxidation and enabled the identification of putative genes for enzymes involved in mitochondrial and peroxisomal beta-oxidation pathways. Several of the genes that showed a specifically altered transcript level in ad7-ACCCA-producing cultures were previously implicated in oxidative stress responses.
Harris
Diana M
DM
Department of Biotechnology, Delft University of Technology, The Netherlands.
Westerlaken
Ilja
I
Schipper
Dick
D
van der Krogt
Zita A
ZA
Gombert
Andreas K
AK
Sutherland
John
J
Raamsdonk
Leonie M
LM
van den Berg
Marco A
MA
Bovenberg
Roel A L
RA
Pronk
Jack T
JT
Daran
Jean-Marc
JM
D000164:Acremonium / Q000502:physiology*; D002511:Cephalosporins / Q000096:biosynthesis*; D024861:Genetic Enhancement / Q000379:methods*; D010408:Penicillium chrysogenum / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D013302:Streptomyces / Q000502:physiology*
D002511:Cephalosporins; D011994:Recombinant Proteins