Formation of pi, tau-dimethylhistidine on alkylation of trypsin with active-site-directed sulfonic acid methyl esters.
The possibility of synthesizing stable alkyl analogues of acyl trypsins by introducing the alkyl residue by means of active-site-directed sulfonic acid esters was studied. Nine amidino- or guanidino-substituted sulfonic acids of different geometries and their methyl esters were prepared. The time-dependent inhibition of bovine trypsin by these esters, indicating modification at the active site of the enzyme, was followed. With the exception of p-guanidinobenzenesulfonic acid methyl ester, all the esters acted as irreversible inhibitors. The site of methylation, Ser-195 or His-57 (chymotrypsinogen numbering), was determine by analyzing for O-methylserine and methylhistidines. With four of the esters indications of a possible formation of, at most, 0.1 residue of O-methylserine per inactivated trypsin molecule were obtained. tau-Methylhistidine (but no pi-methylhistidine) was, however, always observed as the main product of the modification reaction. A further product, hitherto not yet described in active site methylations of serine proteinases, was pi, tau-dimethylhistidine (1,3-dimethylhistidine). The failure of an attempted synthesis of the N-acetyl-ethanolamine ester of p-toluene-sulfonic acid reported in the literature is shown to be due to the high instability of this ester.
Schubert
C
C
Abteilung für Klinische Chemie und Klinische Biochemie, Chirurgische Klinik Innenstadt der Universität, München, Germany.
Fiedler
F
F
D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D004952:Esters / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D008745:Methylation; D008762:Methylhistidines / Q000378:metabolism*; D013451:Sulfonic Acids / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D014357:Trypsin / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*
D000596:Amino Acids; D004952:Esters; D008762:Methylhistidines; D013451:Sulfonic Acids; D014361:Trypsin Inhibitors; C093925:N,N-dimethylhistidine; C028120:1-methylhistidine; D014357:Trypsin