INSIGHT INTO LIPID SURFACE RECOGNITION AND REVERSIBLE CONFORMATIONAL ADAPTATIONS OF AN EXCHANGEABLE APOLIPOPROTEIN BY MULTIDIMENSIONAL HETERONUCLEAR NMR TECHNIQUES

Apolipophorin III (apoLp-III) from the insect Manduca sexta is a 166-r esidue (M-r 18,340) member of the exchangeable apolipoprotein Glass th at functions to stabilize lipid-enriched plasma lipoproteins, Hn the p resent study, we present the secondary structure and global fold of re combinant apoLp-III derived from three-dimensional heteronuclear NMR s pectroscopy experiments. Five discrete alpha-helical segments (21-30 r esidues in length) with well defined boundaries were characterized by four NMR. parameters: medium range nuclear Overhauser enhancement cont acts between proton pairs, chemical shift index, coupling constants, a nd amide, proton exchange rates, An antiparallel arrangement of helica l segments has been obtained based an the long range interhelical nucl ear Overhauser enhancement contacts. The NMR solution structure reveal s a globular, up and down helix bundle organization similar to that of Locusta migratoria apoLp-III (Breiter, D. R., Kanost, M. R., Benning, M, M., Wesenberg, G., Law, J. H., Wells, M. A., Rayment, I., and Hold en, H. M. (1991) Biochemistry 30, 603-608). However, a short helix (co mprised of 5 amino, acids) has been identified ill the region between helix 3 and helix 4, This helix is postulated to play a role in lipid surface recognition and/or initiation of binding. Our results also ind icate the existence of buried polar and charged residues in the helix bundle, providing a structural basis for the relatively low stability of apoLp-III in its Lipid-free state. It is suggested that the intrins ic low stability of lipid-free apoLp-III may be important in terms of its ability to undergo a reversible, lipid binding induced, conformati onal change, This study underscores the striking: resemblance in molec ular architecture between insect apoLp-III and the N-terminal domain o f human apolipoprotein E, The potential for application of NMR techniq ues to studies of the exchangeable apolipoprotein, possibly in their b iologically active, lipid-associated state, has broad implications in terms of our understanding of the molecular basis of their physiologic al functions.