Purification and substrate specificity of Staphylococcus hyicus lipase.
The Staphylococcus hyicus lipase gene has been cloned and expressed in Staphylococcus carnosus. From the latter organism the enzyme was secreted into the medium as a protein with an apparent molecular mass of 86 kDa. This protein was purified, and the amino-terminal sequence showed that the primary gene product was indeed cleaved at the proposed signal peptide cleavage site. The protein was purified from large-scale preparations after tryptic digestion. This limited proteolysis reduced the molecular mass to 46 kDa and increased the specific activity about 3-fold. Although the enzyme had a low specific activity in the absence of divalent cations, the activity increased about 40-fold in the presence of Sr2+ or Ca2+ ions. The purified lipase has a broad substrate specificity. The acyl chains were removed from the primary and secondary positions of natural neutral glycerides and from a variety of synthetic glyceride analogues. Thus triglycerides were fully hydrolyzed to free fatty acid and glycerol. The enzyme hydrolyzed naturally occurring phosphatidylcholines, their synthetic short-chain analogues, and lysophospholipids to free fatty acids and water-soluble products. The enzyme had a 2-fold higher activity on micelles of short-chain D-lecithins than on micelles composed of the L-isomers. Thus the enzyme from S. hyicus has lipase activity and also high phospholipase A and lysophospholipase activity.
van Oort
M G
MG
Department of Biochemistry, State University of Utrecht, The Netherlands.
Deveer
A M
AM
Dijkman
R
R
Tjeenk
M L
ML
Verheij
H M
HM
de Haas
G H
GH
Wenzig
E
E
Götz
F
F
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D003001:Cloning, Molecular; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008049:Lipase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D050356:Lipid Metabolism; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010194:Pancreatin / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D013210:Staphylococcus / Q000201:enzymology* / Q000235:genetics; D013324:Strontium / Q000494:pharmacology; D013379:Substrate Specificity; D013552:Swine; D014280:Triglycerides / Q000494:pharmacology
D010743:Phospholipids; D014280:Triglycerides; D010194:Pancreatin; D008049:Lipase; C005830:tributyrin; D002118:Calcium; D013324:Strontium