Subcellular distribution of lysophospholipase of rat intestinal mucosa.
We have studied the subcellular localization of rat intestinal lysophospholipase activity and some of the biochemical properties of this enzyme. After subcellular fractionation, an enriched activity was found in the high-speed pellet fraction containing the microsomes and the brush border membranes. Subsequently, these organelles were isolated. Using the classical calcium-precipitation method to isolate brush border membranes, we failed to demonstrate any significant recovery of lysophospholipase activity associated with this fraction. The microsomal fraction was further isolated after density gradient centrifugation, and most of the lysophospholipase activity was recovered with this fraction. Because further purification of the enzyme was unsuccessful, some of the biochemical properties of the enzyme were determined on the partially purified microsomal fraction. The optimum pH of the activity was centered at 7.0, and the enzyme did not require bivalent cations. By using double reciprocal plots, we determined the Kapp(m) to be 0.4 mM; the Vapp(max), 23 mumol.h-1.mg protein-1. The enzyme was strongly inhibited by detergents having a low critical micellar concentration and less inhibited by those having a higher critical micellar concentration.
Charbonnier
M
M
Institut national de la santé et de la recherche médicale, Unité 130, Marseille, France.
Grataroli
R
R
Portugal
H
H
Lafont
H
H
Nalbone
G
G
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002413:Cations, Divalent; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D003902:Detergents / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000201:enzymology / Q000648:ultrastructure*; D007700:Kinetics; D008245:Lysophospholipase / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008345:Manganese / Q000494:pharmacology; D008861:Microsomes / Q000201:enzymology; D008871:Microvilli / Q000201:enzymology; D010740:Phospholipases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D002413:Cations, Divalent; D003902:Detergents; D008345:Manganese; D010740:Phospholipases; D008245:Lysophospholipase; D008274:Magnesium; D002118:Calcium