Purification and characterization of LasD: a second staphylolytic proteinase produced by Pseudomonas aeruginosa.
We have previously described studies of a 22 kDa active fragment of the LasA proteinase. In follow-up studies of LasA, we have discovered the separate existence of a 23 kDa proteinase which shares many of the enzymatic properties of LasA, including the ability to lyse heat-killed staphylococci. However, this apparent serine proteinase, which we designate LasD, is distinct from the 22 kDa active LasA protein for the following reasons: (i) the N-terminal sequence of LasD shares no homology with LasA or the LasA precursor sequence; (ii) Pseudomonas aeruginosa LasA mutant strains AD1825 and FRD2128 do not produce LasA yet produce LasD; and (iii) specific antibodies to each proteinase do not show any cross-reactivity. LasD appears to be produced as a 30 kDa protein, which is possibly cleaved to produce a 23 kDa active fragment. The purified LasD fragment (23 kDa) shows strong staphylolytic activity only at higher pH conditions, while LasA exhibits staphylolytic activity over a broad pH range. In addition to their ability to cleave at internal diglycine sites, both the LasD and LasA endoproteinases efficiently cleave beta-casein.
Park
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Department of Microbiology, Ohio State University, Columbus 43210-1292, USA.
Galloway
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D002364:Caseins / Q000378:metabolism; D010450:Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism; D007700:Kinetics; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D008969:Molecular Sequence Data; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000378:metabolism*; D017385:Sequence Homology; D012697:Serine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000378:metabolism; D013379:Substrate Specificity
D001426:Bacterial Proteins; D002364:Caseins; D004791:Enzyme Inhibitors; D015842:Serine Proteinase Inhibitors; D010450:Endopeptidases; C095769:LasD protein, Pseudomonas aeruginosa; D012697:Serine Endopeptidases; D008666:Metalloendopeptidases; C066371:staphylolytic protease