Ivery T C, Daron H H, Aull J L
J Inorg Biochem. 1984 Dec;22(4):259-70. doi: 10.1016/0162-0134(84)85011-4.
Thymidylate synthase from methotrexate-resistant Lactobacillus casei rapidly lost about 90% of its catalytic activity when incubated with an equimolar concentration of IO4- at 0 degree C. Nearly complete inhibition resulted when the IO4- concentration was twice the enzyme concentration or higher. The inhibition reaction appeared to be pseudo-first-order with respect to enzyme when IO4- was in excess. The substrate dUMP, the product dTMP, and inorganic phosphate all protected the enzyme from inactivation by IO4-, with the order of effectiveness: dUMP greater than dTMP greater than phosphate. Deoxyuridine, which is not a substrate, did not protect the enzyme. Titrations with dithiobis(2-nitrobenzoate) (DTNB) showed that approximately 1.5 titratable SH groups were lost when thymidylate synthase was completely inhibited by IO4-. Essentially no reactivation occurred when periodate-inhibited enzyme was dialyzed against buffered 2-mercaptoethanol (ME) or dithiothreitol (DTT). Enzyme that had been treated with p-hydroxymercuribenzoate, DTNB, or methylmethanethiosulfonate prior to treatment with periodate could be completely reactivated with ME or DTT.
