JBC, Vol. 250, Issue 17, 6666-6671, Sep, 1975
Molecular aspects of the inactivation of tryptophanyl transfer ribonucleic acid synthetase by N-ethylmaleimide
F. Iborra, C. Gros, B. Labouesse and J. Labouesse
The tryptic maps of tryptophanyl-tRNA synthetase from beef pancreas show
that the 8 cysteinyl residues of the enzyme subunit are located, 2 by 2, on
four different peptides. The kinetics of the incorporation of radioactivity
from N-[ethyl-14C]ethylmaleimide into these peptides are compared in this
paper with the kinetics of the changes of the catalytic properties of the
enzyme occurring during alkylation. This comparison allows the
identification of (a) the peptide carrying the cysteinyl residues located
on the surface of the molecule, (b) the peptide carrying the deeply buried
residues unmasked by the dissociation of the subunits, and (c) the peptide
carrying the --SH group located in the vicinity of the binding site of
tryptophan. The fourth peptide is shown to have a great sensitivity to pH
with respect to the reactivity of its cysteinyl residues toward
N-ethylmaleimide. The same unusual pH dependence is found for the rate of
quenching of the intrinsic fluorescence of the protein during the
alkylation, suggesting a strong sensitivity of the conformation of
tryptophanyl-tRNA synthetase to pH in the range of 7 to 9.
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