Studies on the Molecular Mechanism of Mersalyl and 4-Aminophenylmercuric Acetate Re-activation of Trypsin-Thiol Complexes

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Abstract
Trypsin was reacted with dithiothreitol and with a naturally occurring thiol-containing trypsin inhibitor to form enzyme-inhibitor complexes. This complex formation is known to be via a reversible intermolecular disulfide linkage. These latent forms of trypsin were re-activated with mersalyl [N-(O-carboxymethylsalicyloyl)-3-hydroxymercuric-2-methoxypropylamine], 4-aminophenylmercuric acetate and with cystine. Active-site titration analysis of trypsin in the presence of incremental additions of dithiothreitol demonstrated the simultaneous inhibition and modification of the enzyme active site, demonstrating a direct involvement of a significant disulfide controlling the conformation of the active site of the enzyme. Mersalyl addition to the dithiothreitol-reduced trypsin resulted in a regain of enzymic activity and a corresponding regain of availability of the active sites for titration. Mersalyl and 4-aminophenylmercuric acetate re-activated the trypsin-inhibitor complex. A molecular mechanism for the organomercurial re-activation of latent enzymes of this particular type (involving disulfide exchange) is proposed.