Thiol ester role in correct folding and conformation of human α2‐macroglobulin

FEBS Letters

Gettins, Peter G.W.²  Crews, Brenda C.²  Boel, Esper¹ 

[1] Department of Molecular Biology, Pharmaceutical Biotechnology, Novo Nordisk AIS. DK-2880 Bagsvard, Denmark;Department of Biochemistry and Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA

关键词: α2-Macroglobulin;    Thiol ester;    Site-directed mutagenesis;    Baby hamster kidney cell;    Human;    α2M;    α2-macroglobulin;    BHK;    baby hamster kidney;    HNE;    human neutrophil elastase;    SDS;    sodium dodecyl sulfate;    PAGE;    polyacrylamide gel electrophoresis;    DMEM;    Dulbecco's modified Eagle's medium;   

DOI  :  10.1016/0014-5793(94)80430-3

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

To determine the role of the thiol ester in the folding of human α₂-macroglobulin (α₂M) in the active conformation, we have characterized a recombinant variant of α₂M, C949S, expressed in baby hamster kidney cells, that lacks the thiol ester-forming cysteine. C949S α₂M behaves like methylamine-treated plasma α₂M, with correctly formed inter-subunit disulfide bridges, non-covalent association of covalent dimers to form tetramers, and exposure of the receptor binding domain, but an inability to inhibit proteinases, and inaccessibility of the bait regions to proteolysis. We concluded that correct folding of monomers or their association to give tetrameric α₂M does not require a pre-formed thiol ester. Active α₂M may form in vivo by a two-step process involving initial folding to give a structure resembling that of C949S α₂M followed by thiol ester formation and a conformational change that gives the native active state.

【 授权许可】

Unknown