Effects of amino group modification in discoidal apolipoprotein A-I-egg phosphatidylcholine-cholesterol complexes on their reactions with lecithin:cholesterol acyltransferase.
Discoidal complexes of human apolipoprotein A-I-egg phosphatidylcholine-cholesterol were prepared by the sodium cholate dialysis procedure and were reacted to varying extents with the amino group reagents citraconic anhydride, diketene, and formaldehyde in the presence of sodium borohydride. Modification of positive lysine residues with negative or neutral groups (citraconic anhydride and diketene, respectively) resulted, for extensively reacted complexes (90%), in structural alterations and in a marked decrease in reactivity with purified human lecithin:cholesterol acyltransferase. The structural and kinetic effects were partially reversible by removal of the modifying groups or by increased ionic strength. Similar extents of modification (84%) with retention of positive charge and introduction of two methyl groups (reductive methylation) had no effect on the structure or the reactivity of the complexes. These results, together with kinetic data at variable complex concentrations or at variable temperatures, indicate that specific lysine residues of apolipoprotein A-I are not involved in the lecithin:cholesterol acyltransferase activation process; instead, charge interactions and structural changes are responsible for the observed decrease in activating capacity. In terms of kinetic parameters, intrinsic K*m values and probably enzyme-substrate particle dissociation constants are affected, but the activation energies remain the same upon chemical modification.
D016632:Apolipoprotein A-I; D001054:Apolipoproteins A / Q000097:blood*; D001894:Borohydrides / Q000494:pharmacology; D002250:Carbon Radioisotopes; D002784:Cholesterol / Q000378:metabolism*; D002949:Citraconic Anhydrides / Q000494:pharmacology*; D004530:Egg Yolk; D005557:Formaldehyde / Q000494:pharmacology*; D005663:Furans / Q000494:pharmacology*; D006801:Humans; D007700:Kinetics; D007783:Lactones / Q000494:pharmacology*; D008075:Lipoproteins, HDL / Q000097:blood*; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000378:metabolism*; D010713:Phosphatidylcholines / Q000378:metabolism*; D011485:Protein Binding
D016632:Apolipoprotein A-I; D001054:Apolipoproteins A; D001894:Borohydrides; D002250:Carbon Radioisotopes; D002949:Citraconic Anhydrides; D005663:Furans; D007783:Lactones; D008075:Lipoproteins, HDL; D010713:Phosphatidylcholines; C004773:diketene; D005557:Formaldehyde; D002784:Cholesterol; D007862:Phosphatidylcholine-Sterol O-Acyltransferase