Specific covalent modification of the tryptophan residues in murine interleukin-6. Effect on biological activity and conformational stability.


Abstract

Modification of recombinant murine interleukin-6 (mIL-6) with the tryptophan-specific reagent 2-nitrophenylsulfenyl chloride under mild acidic conditions, 0.1 M sodium acetate, pH 3.5, yielded a derivative containing 2.02 mol 2-nitrophenylsulfenyl tryptophan/mol protein. The sites of modification were identified as Trp36 and Trp160. No detectable side reactions occurred on other amino acids in the molecule, as indicated by the combination of endoproteinase Asp-N peptide mapping, Edman degradation and electrospray mass spectrometry. Sulfenylation of the two tryptophan residues in mIL-6 caused a 50% reduction in both the biological activity in the murine-hybridoma-growth-factor assay using 7TD1 cells and receptor-binding affinity to mIL-6 receptors. Sulfenylation of mIL-6 did not significantly affect the overall conformation of the protein as measured by farultraviolet circular dichroism and binding to the neutralizing anti-mIL-6 mAb 6B4. The sulfenylated protein was, however, significantly less stable [delta delta G(H2O) = 3.98 kJ/mol] than unmodified mIL-6 as measured by urea-gradient gel electrophoresis. Study holds ProTherm entries: 10791 Extra Details: tryptophan-specific reagent; sulfenylation; biological activity;,receptor-binding affinity; conformation

Submission Details

ID: tuFp2KHz3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:40 p.m.

Version: 1

Publication Details
Zhang JG;Reid GE;Moritz RL;Ward LD;Simpson RJ,Eur. J. Biochem. (1993) Specific covalent modification of the tryptophan residues in murine interleukin-6. Effect on biological activity and conformational stability. PMID:8223586
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