Role of the gene encoding lipase activator from Pseudomonas sp. strain KWI-56 in in vitro activation of lipase.
The function of the gene, act, which encodes lipase activator and that is downstream from the gene that encodes lipase, lip, in Pseudomonas sp. strain KWI-56, was studied with Escherichia coli as the host organism. E. coli carrying both the lip and act genes in either cis or trans produced an active lipase, but E. coli carrying only the lip gene produced an equal amount of inactive lipase protein. The active and inactive lipases had the same molecular weight and the same N-terminal amino acid sequence, so their primary structures were the same. The inactive lipase was not activated spontaneously, but it was activated in vitro by the addition of a crude cell extract of E. coli carrying an expression plasmid of the act gene. Lipase from Pseudomonas sp. strain KWI-56 was completely without enzymatic activity when denatured, but regained activity in vitro upon addition of the crude cell extract. These results indicate that the act gene affects the activation involved in the conformational change of the lipase protein to its active form.
Iizumi
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Kurita Central Laboratories, Kurita Water Industries Ltd., Atsugi, Japan.
Fukase
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D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000378:metabolism; D004789:Enzyme Activation; D004926:Escherichia coli; D005798:Genes, Bacterial; D007700:Kinetics; D008049:Lipase / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010957:Plasmids; D011489:Protein Denaturation; D011549:Pseudomonas / Q000201:enzymology / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D015183:Restriction Mapping; D017384:Sequence Deletion; D015534:Trans-Activators / Q000096:biosynthesis / Q000235:genetics*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D015534:Trans-Activators; C088618:lipase activator protein, Bacteria; D008049:Lipase