Structure and Stability of Apolipoprotein A-I in Solution and in Discoidal High-Density Lipoprotein Probed by Double Charge Ablation and Deletion Mutation
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- 作者：Irina N. Gorshkova ; Tong Liu ; Horng-Yuan Kan ; Angeliki Chroni ; Vassilis I. Zannis ; and David Atkinson
- 刊名：Biochemistry
- 出版年：2006
- 出版时间：January 31, 2006
- 年：2006
- 卷：45
- 期：4
- 页码：1242 - 1254
- 全文大小：325K
- 年卷期：v.45,no.4(January 31, 2006)
- ISSN：1520-4995
文摘
To identify residues and segments in the central region of apolipoprotein A-I (apoA-I) thatare important for the protein structure and stability, we studied the effects of four double charge ablations, D102A/D103A, E110A/E111A, R116V/K118A, and R160V/H162A, and two deletion mutations,
(61-78) and (121-142), on the conformation and stability of apoA-I in the lipid-free state and inreconstituted discoidal phospholipid-cholesterol-apoA-I particles (rHDL). The findings suggest thatD102/D103 and E110/E111 located in helix 4 and segment(s) between residues 61 and 78 are involvedin maintenance of the conformation and stability of apoA-I in both the lipid-free state and in rHDL.R116/K118 located in helix 4 are essential for the conformation and stabilization of apoA-I in rHDL butnot vital for the lipid-free state of the protein. The R160V/H162A substitutions in helix 6 lead to a lesscompact tertiary structure of lipid-free apoA-I without notable effects on the lipid-free or lipid-boundsecondary conformation, suggesting involvement of R160/H162 in important interhelical interactions.The results on the (121-142) mutant, together with our earlier findings, suggest disordered structure ofa major segment between residues 121 and 143, likely including residues 131-143, in lipid-free apoA-I.Our findings provide the first experimental evidence for stabilization of rHDL by specific electrostaticinterhelical interactions, in agreement with the double belt model. The effects of alterations in theconformation and stability of the apoA-I mutants on in vitro and in vivo functions of apoA-I and lipidhomeostasis are discussed.
(61-78) and (121-142), on the conformation and stability of apoA-I in the lipid-free state and inreconstituted discoidal phospholipid-cholesterol-apoA-I particles (rHDL). The findings suggest thatD102/D103 and E110/E111 located in helix 4 and segment(s) between residues 61 and 78 are involvedin maintenance of the conformation and stability of apoA-I in both the lipid-free state and in rHDL.R116/K118 located in helix 4 are essential for the conformation and stabilization of apoA-I in rHDL butnot vital for the lipid-free state of the protein. The R160V/H162A substitutions in helix 6 lead to a lesscompact tertiary structure of lipid-free apoA-I without notable effects on the lipid-free or lipid-boundsecondary conformation, suggesting involvement of R160/H162 in important interhelical interactions.The results on the (121-142) mutant, together with our earlier findings, suggest disordered structure ofa major segment between residues 121 and 143, likely including residues 131-143, in lipid-free apoA-I.Our findings provide the first experimental evidence for stabilization of rHDL by specific electrostaticinterhelical interactions, in agreement with the double belt model. The effects of alterations in theconformation and stability of the apoA-I mutants on in vitro and in vivo functions of apoA-I and lipidhomeostasis are discussed.