NEW CONFORMATIONAL PROPERTIES INDUCED BY THE REPLACEMENT OF TYR-64 INDESULFOVIBRIO-VULGARIS HILDENBOROUGH FERRICYTOCHROME C(553) USING ISOTOPIC EXCHANGES MONITORED BY MASS-SPECTROMETRY

In order to study the conformational stability induced by the replacem ent of Tyr-64 in Desulfovibrio vulgaris Hildenborough (DvH) cytochrome c(553), fast peptic digestion of deuterated protein followed by separ ation and measurement of related peptides using liquid chromatography coupled to electrospray ionization mass spectrometry was performed, We show that the H-bonding and/or solvent accessibility properties mere modified by the single-site mutation, The mutant proteins can be class ified into two groups: the Y64F and Y64L mutants with nearly unchanged deuterium incorporation compared to the wildtype protein and the Y64S , Y64V and Y64A mutants with increased deuterium incorporation, The 70 -74 peptide was the most affected by mutation of Tyr-64, the phenylala nine mutant inducing slight stabilization whereas the serine mutant ma s significantly destabilized. In addition, from the analysis of the ov erlapping 37-57 and 38-57 peptides we can conclude that the amide prot on of Tyr-38 has been replaced by deuterium in all proteins.