Histidyl-transfer-ribonucleic-acid synthetase from Salmonella typhimurium. Studies of the sulfhydryl groups

The reactivity of the SH groups of histidyl-tRNA synthetase (I) of S. typhimurium and the effect of substrates on the reactivity was studied using p-hydroxymercuribenzoate (II) and 5,5'-dithiobis(2-nitrobenzoic acid) (III) as reagents. III reacted with only 2 SH groups/I mol. and the reaction was essentially monophasic, while II reacted with 4 SH groups. As obsd. kinetically the reaction with II is strongly biphasic, each phase corresponding to ∼2 SH groups/I mol. With both reagents, no detectable difference in SH group reactivity was obsd. when ATP, histidine, and tRNAHis were added individually or in combination to I. I activity slowly changed after 2 or 4 SH groups were blocked by III or II, resp. A new, stable level of activity was reached that was characterized by a different Km value for the aminoacylation reaction. The results indicate that the SH groups reacting with the 2 reagents used here are not directly involved either in the binding of the substrates or in the catalytic process. The ultimate change in I activity after reaction of the SH groups suggests a transition to an alternative structure. [on SciFinder(R)]