Single repeat deletion in ApoA-I blocks cholesterol esterification and results in rapid catabolism of Δ6 and wild-type ApoA-I in transgenic mice

Abstract

The deletion mutation Δ6 apolipoprotein A-I lacks residues 143-164 or repeat 6 in the mature apoA-I protein. In vitro studies show this mutation dramatically reduces the rate of lecithin:cholesterol acyltransferase (LCAT) catalyzed cholesterol esterification. The present study was initiated to investigate the effect of this mutation on in vivo high density lipoprotein (HDL) cholesterol esterification and metabolism. Transgenic mice expressing human Δ6 apoA-I (TgA6 +/+) were created and then crossed with apoA-I knockout mice (-/-) to generate mice expressing only human Δ6 apoA-I (TgΔ6 -/-). Human Δ6 apoA-I was associated with homogeneous sized α-HDL, when wild-type mouse apoA-I was present (in TgΔ6 +/+ and +/- mice). However, in the absence of endogenous mouse apoA-I, Δ6 apoA-I was found exclusively in cholesterol ester-poor HDL, and lipid-free HDL fractions. This observation coincides with the 6-fold lower cholesterol ester mass in TgΔ6 -/- mouse plasma compared with control. Structural studies show that despite the structural perturbation of a domain extending from repeat 5 to repeat 8 (137- 178), Δ6 apoA-I binds to spherical unilamellar vesicles with only 2-fold less binding affinity. In summary, these data show a domain corresponding to apoA-I repeat 6 is responsible for providing an essential conformation for LCAT catalyzed generation of cholesterol esters. Deletion of apoA-I repeat 6 not only blocks normal levels of cholesterol esterification but also exerts a dominant inhibition on the ability of wild-type apoA-I to activate LCAT in vivo.