Mapping the mechanism-based modification sites in L-aspartase from Escherichia coli

Archives of Biochemistry and Biophysics
F GiorgianniR E Viola

Abstract

Inactivation of the enzyme L-aspartase from Escherichia coli by the substrate analog aspartate beta-semialdehyde has previously been shown to occur by the mechanism-based conversion to the corresponding product aldehyde, followed by covalent modification of cysteine-273 (F. Giorgianni et al. (1995) Biochemistry 34, 3529). Inactivation by the product analog, fumaric acid aldehyde (FAA), has now been examined directly by adding a reduction step to the modification protocol in order to stabilize the resulting enzyme-FAA derivative(s). HPLC and mass spectrometric analyses of proteolytic digests of inactivated L-aspartase have confirmed the modification at cysteine-273, and have also identified an additional modified peptide. The inactivation at this additional site involves a crosslink between cysteine-140 and an adjacent lysine. Site-directed mutagenesis studies have shown that cysteine-140 is a very reactive and accessible nucleophile that is not, however, directly involved in enzyme activity. The adjacent lysine-139 that is modified does appear to play a role in substrate binding. A double mutant in which both of the reactive cysteines have been replaced is almost completely insensitive to modification by these substrate and pro...Continue Reading

References

Sep 22, 1975·Biochimica Et Biophysica Acta·K Mizuta, M Tokushige
Jan 1, 1979·Methods in Enzymology·W W Cleland
Jan 1, 1977·Methods in Enzymology·G R Drapeau
May 15, 1991·Archives of Biochemistry and Biophysics·W E Karsten, R E Viola
Jan 1, 1991·Methods in Enzymology·T A KunkelJ McClary
May 31, 1991·Biochemical and Biophysical Research Communications·S MuraseM Tokushige
Jul 31, 1986·Biochemical and Biophysical Research Communications·J S TakagiM Kanehisa
Dec 15, 1983·European Journal of Biochemistry·K PalczewskiM Kochman
Feb 11, 1982·Biochemical and Biophysical Research Communications·N YumotoM Tokushige
Aug 1, 1995·Nature Structural Biology·T M WeaverL J Banaszak
Jan 1, 1995·Methods in Enzymology·R B Silverman
Aug 9, 1994·Biochemistry·J F Schindler, R E Viola
Dec 20, 1993·Journal of Molecular Biology·W ShiG K Farber

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