MOLECULAR-GENETICS OF THE EXTRACELLULAR LIPASE OF PSEUDOMONAS-AERUGINOSA PAO1

The structural gene (lip A) coding for the extracellular lipase of Pseudomonas aeruginosa PAO1 has been cloned on plasmid pSW118. Nucleotide sequence analysis revealed a gene of 936 bp. lip A codes for a proenzyme of 311 amino acids including a leader sequence of 26 amino acids. The mature protein was predicted to have a M(r) of 30134, an isoelectric point of 5.6, and a consensus sequence (IGHSHGG) typical of lipases. Furthermore it is highly homologous (> 60%) to other lipases from various pseudomonads. The lip A gene failed to hybridize detectably with genomic DNA from other Pseudomonas species except P. alcaligenes, even under relaxed stringency. Located 220 bp downstream of the lip A gene, is an open reading frame (ORF2, lip H) which encodes a hydrophilic protein (283 amino acids; M(r) 33587) that shows some homology to the limA gene product of P. cepacia. In complementation tests of lipase-defective mutants, lip H was shown to be necessary for expression of active extracellular lipase in P. aeruginosa PAO1.