Apolipoprotein A-I domains involved in the activation of lecithin:cholesterol acyltransferase. Importance of the central domain.
The reaction of highly purified lecithin:cholesterol acyltransferase (LCAT) with defined reconstituted discoidal apoA-I-containing lipoproteins (LpA-I) with 2, 3, or 4 apoA-I molecules/particle (Lp2, 3, or 4A-I) has been studied in the presence of a number of specific anti apoA-I antibodies. Among nine anti-apoA-I monoclonal antibodies (mAbs) reacting with epitopes distributed over 80% of the sequence, three significantly inhibit the LCAT reaction with all particles. The position of their epitopes located in the middle to COOH-terminal region between residues 96-121 (3G10), 135-148 (A03), and 149-186 (A44) is compatible with an inhibition by steric hindrance over a central domain. Antibody 4H1 binding to the NH2 terminus (residues 2-8) profoundly increases (5-fold) the LCAT reaction with Lp2A-I (7.8 nm), but not with other particles. Other mAbs, A11 and 5F6, binding to epitopes (residues 99-139 and 118-141) enhance LCAT reactivity with the small Lp2A-I (7.8 nm) and Lp3A-I (10.8 nm) but not with their larger counterparts. Most mAbs have similar effects on LCAT reaction with native high density lipoprotein3 as with LpA-I. The inhibitory or enhancing effects of these mAbs are also observed with Fab fragments and not related to their binding affinity for apoA-I containing reconstituted lipoprotein particles. The intercalation of epitopes for mAbs that inhibit or enhance LCAT reaction with small LpA-I is compatible not with steric hindrance but with conformational modifications of apoA-I and indirectly of the lipids in small particles. We propose that enhancing mAbs act by stabilization of an apoA-I conformation which is not favored in small LpA-I, i.e. by increasing binding of amphipathic helices to lipids or by interfering with the mobility of a hinged domain. The epitopes for the inhibitory mAbs can be shown to overlap on several LpA-I models, indicating that steric hindrance over a single site is a possible mechanism of inhibition.
Meng
Q H
QH
Lipoproteins and Atherosclerosis Group, University of Ottawa Heart Institute, Ontario, Canada.
Calabresi
L
L
Fruchart
J C
JC
Marcel
Y L
YL
D000911:Antibodies, Monoclonal; D016632:Apolipoprotein A-I / Q000378:metabolism*; D004789:Enzyme Activation; D000939:Epitopes; D006801:Humans; D007862:Phosphatidylcholine-Sterol O-Acyltransferase / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D011233:Precipitin Tests
D000911:Antibodies, Monoclonal; D016632:Apolipoprotein A-I; D000939:Epitopes; D007862:Phosphatidylcholine-Sterol O-Acyltransferase