Kinetic characterization of alkaline mesentericopeptidase. Comparison with serine proteinases from different origins.
Comparative studies of the hydrolysis of succinyl-Ala2-Phe-methylcoumarylamide with mesentericopeptidase, a mesophilic extracellular serine proteinase from Bacillus mesentericus, and proteinases produced by organisms representing different levels of evolutionary development, were performed. Drastic differences in the proteolytic coefficient kcat/Km were found. As regards their catalytic efficiency, the proteinases studied can be placed in the following order: mesentericopeptidase < subtilisin Novo << subtilisin DY < proteinase K < subtilisin Carlsberg < thermitase < alpha-chymotrypsin. The size of the substrate-binding site of mesentericopeptidase for synthetic peptides was studied by using chloromethyl ketones with the general formula benzyloxycarbonyl-Alan-Phe-CH2Cl (n = 1, 2, 3). The presence of at least five binding subsites (S1 ... S5) on the S-side of the hydrolysed bond was suggested. Studies of the primary specificity of mesentericopeptidase with a series of dipeptide chloromethyl ketones having the general formula benzyloxycarbonyl-Ala-Aa-CH2Cl (Aa = Ala, Val, Leu, Phe) revealed the following order of reactivity toward these inhibitors: Aa = Leu >> Ala > Phe > Val. Kinetically, mesentericopeptidase is similar to subtilisin BPN'/Novo.
Dolaschka
P
P
Institute of Organic Chemistry, Bulgarian Academy of Sciences, Sofia.
Genov
N
N
Ermer
A
A
Peters
K
K
Fittkau
S
S
D004151:Dipeptides / Q000494:pharmacology*; D007700:Kinetics; D012697:Serine Endopeptidases / Q000187:drug effects* / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*
D004151:Dipeptides; D015842:Serine Proteinase Inhibitors; D012697:Serine Endopeptidases; C021553:mesentericopeptidase