Structural modulation of salt-resistant rat-liver lipase alters the relative phospholipase and triacylglycerol hydrolase activities.
This paper demonstrates that structural modification of the heparin-releasable salt-resistant lipase of rat liver (liver lipase) alters its relative capacity to hydrolyze phospholipid and triacylglycerol emulsions. Enzymatic activities were modified by immunoinhibition and proteolysis and by selective amino acid agents. Binding of three different monoclonal antibodies resulted in a lower extent of inhibition of phospholipase than of triacylglycerol hydrolase activity. Degradation of the enzyme by trypsin under mild conditions led to a decrease of both enzyme activities in a different way. Triacylglycerol hydrolase activity was less affected than the phospholipase activity. Visualization of the proteolysis of the purified enzyme by immunoblotting revealed the actual breakdown of a 58 kDa protein into a 53 kDa protein band and subsequently in a 48 kDa one. Incubation of the purified enzyme by N-tosyl-L-phenylethylchloromethyl ketone (acting on cysteine or histidine) or N-ethylmaleimide (a sulfhydryl reagent) did not influence either enzyme activity. On the other hand, after the selective modification of lysine residue(s) by phenylisothiocyanate, the phospholipase A1 activity was stimulated by 68%, whereas the triacylglycerol hydrolase activity was completely lost. The role of a lysine residue(s) in the activity of the enzyme towards phospholipid and triacylglycerol emulsions is discussed.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D017879:Isothiocyanates; D007700:Kinetics; D008049:Lipase / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054466:Phospholipases A1; D051381:Rats; D012492:Salts; D013861:Thiocyanates / Q000494:pharmacology; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology; D014357:Trypsin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D017879:Isothiocyanates; D012492:Salts; D013861:Thiocyanates; C005441:phenylisothiocyanate; D014108:Tosylphenylalanyl Chloromethyl Ketone; D010740:Phospholipases; D008049:Lipase; D010741:Phospholipases A; D054466:Phospholipases A1; D014357:Trypsin