To investigate the structure-function relationship of human apolipoprotein A-IV (apoA-IV), several deletion mutants of this protein were constructed by sequentially removing pairs of 22-residue repeats, potentially having an amphipathic alpha-helical conformation. The mutants, produced as recombinant poly-histidine-tagged apolipoproteins (t-apo) in Escherichia coli, assembled with phosphatidylcholine (i.e. dimyristoylphosphatidylcholine, palmitoyloleoylphosphatidylcholine, or egg lecithin) as did native apoA-IV. Lecithin:cholesterol acyltransferase (LCAT) cofactor function, measured as cholesterol esterification occurring when 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%, respectively, 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.