Interconversion of D-ribose-5-phosphate (R5P) and D-ribulose-5-phosphate is an important step in the pentose phosphate pathway. Two unrelated
enzymes with R5P
isomerase activity were first identified in Escherichia coli, RpiA and RpiB. In this organism, the essential 5-carbon
sugars were thought to be processed by RpiA, while the primary role of RpiB was suggested to instead be interconversion of the rare 6-carbon
sugars D-allose-6-phosphate (All6P) and D-allulose-6-phosphate. In Mycobacterium tuberculosis, where only an RpiB is found, the 5-carbon
sugars are believed to be the
enzyme's primary substrates. Here, we present kinetic studies examining the All6P
isomerase activity of the RpiBs from these two organisms and show that only the E. coli
enzyme can catalyze the reaction efficiently. All6P instead acts as an inhibitor of the M.
tuberculosis enzyme in its action on R5P. X-ray studies of the M.
tuberculosis enzyme co-crystallized with All6P and 5-deoxy-5-phospho-D-ribonohydroxamate (an inhibitor designed to mimic the 6-
carbon sugar) and comparison with the E. coli
enzyme's structure allowed us to identify differences in the active sites that explain the kinetic results. Two other structures, that of a mutant E. coli RpiB in which
histidine 99 was changed to
asparagine and that of wild-type M.
tuberculosis enzyme, both co-crystallized with the substrate
ribose-5-phosphate, shed additional light on the reaction mechanism of RpiBs generally.