ESSENTIAL HISTIDYL RESIDUES AT THE ACTIVE SITE(S) OF SUCROSE-PHOSPHATE SYNTHASE FROM PROSOPIS-JULIFLORA

Chemical modification of sucrose-phosphate synthase (EC 2.4.1.14) from Prosopis juliflora by diethyl pyrocarbonate (DEP) and photo-oxidation in the presence of rose bengal (RB) which modify the histidyl residue s of the protein resulted in the inactivation of the enzyme activity. This inactivation was dependent on the concentration of the modifying reagent and the time of incubation and followed pseudo-first order kin etics. For both the reagents, the inactivation was maximum at pH 7.5, which is consistent with the involvement and presence of histidine res idues at the active site of the enzyme. Substrates, UDPG and F6P prote cted the enzyme against the inactivation by the modifying reagents sug gesting that the histidine residues may be involved in the binding of these substrates and are essential for the catalytic activity. Specifi city of DEP was indicated by an increase in absorbance at 240 nm along with concomitant inactivation of the enzyme and reactivation of the m odified enzyme by hydroxylamine. These results strongly suggest the pr esence of histidine residue(s) at or near the active site of the enzym e. (C) 1998 Elsevier Science B.V. All rights reserved.