Pepsin inhibition by a high specific activity radioiodinated derivative of pepstatin.

Stop-flow kinetic studies are reported for the reaction of glutathione (GSH) with 2-chloromercuri-4-nitrophenol. The second-order rate constant was found to vary by less than twofold in going from pH 5.9 to 7.3, suggesting that the reaction involves attack of the un-ionized glutathione thiol on the mercurial around pH 7. The rate constant was 1.5 × 10⁷m^?1 s^?1 at pH 7.0 and 8 °C in dilute buffer of ionic strength 0.2. The sensitivity of the kinetics to the nature of the leaving group in the mercurial i.e., to the labile ligand, was investigated in complexes of the mercurial with OH^?, N₃^?, Br^?, Cl^?, CN^?, SCN^?, pyrophosphate, ADP, ATP, EDTA, and adenosine. The addition of most of these substances was observed to lead to rate enhancements or rate retardations, the kinetic effects showing typical binding isotherm behavior when plotted as a function of the added ligand concentration. The kinetic midpoints of such plots showed very good correlation with the independently determined affinities they had toward the mercurial. The results demonstrate that many commonly added compounds of biochemical interest can significantly influence mercurial reactivity toward thiols through rapid exchange at the labile ligand position of the mercurial.