Evidence for a hybrid ping pong-semirandom mechanism for human jejunal glutathione disulfide reductase

Initial rate versus substrate concentration. [S], profiles (at pH 7.4, 37 d egreesC) for human jejunal glutathione disulfide reductase indicated partia l substrate inhibition by GSSG at fixed [NADPH] less than or equal to 40 mu M. Saturation by NADPH assumed an increasingly sigmoidal character at fixed [GSSG] greater than or equal to 20 muM. The results were interpreted in te rms of a hybrid ping pong-semirandom mechanism proceeding through both E.NA DPH (ping pong pathway) and E.GSSG (ordered pathway) complexes. When V(appa rent) versus [GSSG] and [NADPH] were plotted: hygerbolic curves were obtain ed. Plotting 1/Vm(apparent) versus 1/[NADPH] and 1/[GSSG] yielded two lines with different slopes but intersecting at the same point on the S axis. A value of 26.3 U/ml was calculated for Vm. and Km values of 25 and 71 muM we re determined for NADPH and GSSG, respectively.