Lipophorin lipase from the yolk of Manduca sexta eggs: identification and partial characterization.
In the hawkmoth Manduca sexta high density lipophorin from adult insects (HDLp-A) delivers lipids to developing oocytes. During this lipid delivery HDLp-A is taken up by the oocyte and converted to a very high density lipophorin (VHDLp), which is stored in protein storage granules (yolk bodies). A membrane-free lysate of isolated M. sexta yolk bodies was demonstrated to contain lipoprotein lipase activity that hydrolyses the diacylglycerol of HDLp-A. With HDLp-A as a substrate yolk body lipophorin lipase (YBLpL) activity was shown to be maximal between pH 9 and pH 9.5. NaCl concentration was optimal between 0.7 M and 1 M. YBLpL activity required neither bovine serum albumin nor calcium ions but appeared to be stimulated by 5 mM EDTA. Diisopropyl fluorophosphate effectively inhibited YBLpL activity, indicating the presence of a serine in the active site of the enzyme. The identified lipase activity co-eluted with lipophorins and vitellins from the yolk in the void volume of a Sephadex G-75 gel filtration column. This observation suggests that the lipase has a Mr of more than 80,000, or that the enzyme is associated with the lipoproteins. Incubation of HDLp-A with yolk body lysate converted HDLp-A to two classes of higher density lipophorins. The highest density lipophorins produced during this incubation approached the density of VHDLp as it is isolated from mature eggs. The possible role of YBLpL activity in the delivery of lipids to developing oocytes is discussed.
Van Antwerpen
R
R
Department of Biochemistry, University of Arizona, Tucson 85721.
Law
J H
JH
D000818:Animals; D002118:Calcium / Q000502:physiology; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D004075:Diglycerides / Q000378:metabolism; D004492:Edetic Acid; D004530:Egg Yolk / Q000201:enzymology* / Q000378:metabolism; D005260:Female; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008049:Lipase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D008074:Lipoproteins; D008075:Lipoproteins, HDL / Q000378:metabolism; D009036:Moths / Q000201:enzymology* / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000502:physiology; D012965:Sodium Chloride / Q000378:metabolism; D013379:Substrate Specificity
D002352:Carrier Proteins; D004075:Diglycerides; D008074:Lipoproteins; D008075:Lipoproteins, HDL; C032830:lipophorin; D012710:Serum Albumin, Bovine; D012965:Sodium Chloride; D004492:Edetic Acid; D008049:Lipase; D002118:Calcium