| Autor: |
Hua QX; Department of Biochemistry, Case Western Reserve School of Medicine, Cleveland, Ohio 44106-4935, USA., Chu YC, Jia W, Phillips NF, Wang RY, Katsoyannis PG, Weiss MA |
| Jazyk: |
angličtina |
| Zdroj: |
The Journal of biological chemistry [J Biol Chem] 2002 Nov 08; Vol. 277 (45), pp. 43443-53. Date of Electronic Publication: 2002 Aug 23. |
| DOI: |
10.1074/jbc.M206107200 |
| Abstrakt: |
The A and B chains of insulin combine to form native disulfide bridges without detectable isomers. The fidelity of chain combination thus recapitulates the folding of proinsulin, a precursor protein in which the two chains are tethered by a disordered connecting peptide. We have recently shown that chain combination is blocked by seemingly conservative substitutions in the C-terminal alpha-helix of the A chain. Such analogs, once formed, nevertheless retain high biological activity. By contrast, we demonstrate here that chain combination is robust to non-conservative substitutions in the N-terminal alpha-helix. Introduction of multiple glycine substitutions into the N-terminal segment of the A chain (residues A1-A5) yields analogs that are less stable than native insulin and essentially without biological activity. (1)H NMR studies of a representative analog lacking invariant side chains Ile(A2) and Val(A3) (A chain sequence GGGEQCCTSICSLYQLENYCN; substitutions are italicized and cysteines are underlined) demonstrate local unfolding of the A1-A5 segment in an otherwise native-like structure. That this and related partial folds retain efficient disulfide pairing suggests that the native N-terminal alpha-helix does not participate in the transition state of the reaction. Implications for the hierarchical folding mechanisms of proinsulin and insulin-like growth factors are discussed. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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