Lipid binding-induced conformational changes in the N-terminal domain of human apolipoprotein E

The N-terminal domain of human apolipoprotein E3 (apoE3) adopts an elongate d, globular four helix bundle conformation in the lipid-free state. Upon li pid binding, the,protein is thought to undergo a significant conformational change that is essential for manifestation of its low density lipoprotein receptor recognition properties. We have used fluorescence resonance energy transfer (FRET) to characterize helix repositioning which accompanies lipi d interaction of this protein, ApoE3(1-183) possesses a single cysteine at position 112 and four tryptophan residues (positions 20, 26, 34, and 39), M odification of Cys112 with the chromophore, N-iodoacetyl-N'-(5-sulfo-1-naph thyl)etheylene- diamine (AEDANS) was specific and did not alter the seconda ry structure content of the protein. The efficiency of energy transfer from donor Trp residues to the AEDANS moiety was 49% in buffer, consistent with close proximity of the chromophores. Guanidine HCl titration experiments i nduced characteristic changes in the efficiency of energy transfer, indicat ing that FRET data faithfully reports on the conformational status of the p rotein. Interaction of AEDANS-apoE3 (1-183) with dimyristoylphosphatidylcho line to form disk particles, or with detergent micelles, resulted in large decreases in the efficiency of energy transfer. Distance calculations based on the FRET measurements revealed that lipid binding increases the average distance between the four Trp donors and the AEDANS acceptor from 23 Angst rom to 44 Angstrom. The results obtained demonstrate the utility of FRET to investigate conformational adaptations of exchangeable apolipoproteins and are consistent with the hypothesis that, upon lipid binding, apoE3(1-183) undergoes conformational opening, repositioning helix 1 and 3 to adopt a re ceptor-active conformation.