The Pseudomonas aeruginosa acsA gene, encoding an acetyl-CoA synthetase, is essential for growth on ethanol

Fachgebiet Technische Biochemie, Institut für Biotechnologie der Technischen Universität Berlin, Seestraße 13, D-13353 Berlin, Germany 1 Author for correspondence: Helmut Görisch. Tel: +49 30 314 27582. Fax: +49 30 314 27581. e-mail: Goerisch{at}lb.TU-Berlin.De Pseudomonas aeruginosa ATCC 17933 uses a pyrroloquinoline quinone-dependent ethanol oxidation system. Two mutants of P. aeruginosa , unable to grow on ethanol and showing no acetyl-CoA synthetase (ACS) activity under standard test conditions, were complemented by cosmid pTB3018. Subcloning led to the isolation of a gene which encodes a protein with high similarity to acetyl-CoA synthetases. Interruption of the putative acsA gene by a kanamycin-resistance cassette resulted in a mutant also unable to grow on ethanol and with very low residual acetyl-CoA-forming activity. Complementation by the wild-type allele of the acsA gene restored growth and led to the expression of ACS activity in excess of that of wild-type cells. In wild-type P. aeruginosa , ACS activity was induced upon growth on ethanol, 2,3-butanediol, malonate and acetate. The wild-type and mutants defective in ACS activity showed an active acetate kinase (ACK) under the growth conditions used; however, phosphotransacetylase (PTA) could not be detected. The data indicate that P. aeruginosa requires active acsA gene product for growth on ethanol. Keywords: ethanol oxidation, acetate metabolism, acetate kinase, phosphotransacetylase, Pseudomonas aeruginosa Abbreviations: ACK, acetate kinase; ACS, acetyl-CoA synthetase; PQQ, pyrroloquinoline quinone; PTA, phosphotransacetylase; QEDH, quinoprotein ethanol dehydrogenase