Apolipoprotein A-I adopts a belt-like orientation in reconstituted high density lipoproteins

Apolipoprotein A-I (apoA-I) is the major protein associated with high densi ty lipoprotein (HDL), and its plasma levels have been correlated with prote ction against atherosclerosis. Unfortunately, the structural basis of this phenomenon is not fully understood. Over 25 years of study have produced tw o general models of apoA-I structure in discoidal HDL complexes. The "belt" model states that the amphipathic helices of apoA-I are aligned perpendicu lar to the acyl chains of the lipid bilayer, whereas the "picket fence" mod el argues that the helices are aligned parallel with the acyl chains. To di stinguish between the two models, various single tryptophan mutants of apoA -I were analyzed in reconstituted, discoidal HDL particles composed of phos pholipids containing nitroxide spin labels at various positions along the a cyl chain. We have previously used this technique to show that the orientat ion of helix 4 of apoA-I is most consistent with the belt model. In this st udy, we performed additional control experiments on helix 4, and we extende d the results by performing the same analysis on the remaining 22-mer helic es (helices 1, 2,5,6,7,8, and 10) of human apoA-I. For each helix, two diff erent mutants were produced that each contained a probe Trp occurring two h elical turns apart. In the belt model, the two Trp residues in each helix s hould exhibit maximal quenching at the same nitroxide group position on the lipid acyl chains. For the picket fence model, maximal quenching should oc cur at two different levels in the bilayer. The results show that the major ity of the helices are in an orientation that is consistent with a belt mod el, because most Trp residues localized to a position about 5 A from the ce nter of the bilayer. This study corroborates a belt hypothesis for the majo rity of the helices of apoA-I in phospholipid discs.