Figure 5.
Reaction of representative active site mutants with protiated and deuterated phosphite in the pre-steady state. A) Reaction of R237K with protiated and deuterated phosphite in the pre-steady state. Reaction conditions: 100 mM MOPS pH 7.25, 100 mM 1H or 2H phosphite, 10 mM NAD+, 33 μM R237K. B) Reaction of E266Q with protiated and deuterated phosphite in the pre-steady state. Reaction conditions: 100 mM MOPS pH 7.25, 100 mM 1H or 2H phosphite, 5 mM NAD+, 15 μM E266Q. Each trace is the average of four separate shots. Background due to absorbance of NAD+ at 340 nm has been subtracted.