IDENTIFICATION OF SPECIFIC AMPHIPATHIC ALPHA-HELICAL SEQUENCE OF HUMAN APOLIPOPROTEIN A-IV INVOLVED IN LECITHIN-CHOLESTEROL ACYLTRANSFERASEACTIVATION

To investigate the structure-function relationship of human apolipopro tein A-IV (apoA-IV), several deletion mutants of this protein were con structed by sequentially removing pairs of ga-residue repeats, potenti ally having an amphipathic cu-helical conformation. The mutants, produ ced as recombinant poly-histidine tagged apolipoproteins (t-apo) in Es cherichia coli, assembled with phosphatidylcholine (i.e. dimyristoylph osphatidylcholine, palmitoyloleoylphosphatidylcholine, or egg lecithin ) as did native apoA-IV. Lecithin:cholesterol acyltransferase (LCAT) c ofactor function, measured as cholesterol esterification occurring whe n t-apo-phosphatidylcholine-cholesterol complexes were incubated with purified enzyme, decreased significantly when pairs of repeats between residues 117 and 248 were deleted and most markedly when residues 117 -160 were deleted. LCAT cofactor activity decreased by 90 and 75%, res pectively, when egg lecithin or palmitoyloleoylphosphatidylcholine was used to form the particles with the Delta aa 117-160 mutant, Thus, on the basis of deletion scanning of t-apo, residues 117-160 seem to be involved in the LCAT cofactor function of apoA-IV.