Thymidine Phosphorylase from Escherichia coli. Properties and Kinetics

Thymidine phosphorylase has been purified several hundred-fold from a strain of Escherichia coli K 12. The molecular weight of the enzyme was determined to be 68000 ± 10% both by use of G-200 gel filtration and by use of dodecylsulphate gel electrophoresis. The purified enzyme has been subjected to kinetic analyses, by the method described by Cleland, including initial velocities, base-base exchange of the deoxyribosyl group, and product inhibition studies. The results are consistent with an ordered sequential mechanism in which phosphate is the first substrate to add to, and deoxyribose 1-phosphate is the last substrate to leave the enzyme.