Substrate and inhibitor specificity of glutamine cyclotransferase (QC).

This paper reports a systematic study of the substrate and inhibitor specificity of papaya latex glutamine cyclotransferase (QC). The results showed that the second amino acid residue in N-terminal glutaminyl peptides significantly accelerated papaya latex QC-catalyzed reactions while the third residue provided no further rate enhancement. Substrate binding was shown to be the main specificity-determining step.

The second nucleophile molecule binds to the acyl-enzyme-nucleophile complex in alpha-chymotrypsin catalysis. Kinetic evidence for the interaction.

alpha-Chymotrypsin-catalyzed acyl transfer was studied using three acyl-group donors (Mal-L-Ala-L-Ala-L-PheOMe, Bz-L-TyrOEt and Ac-L-TrpOEt; Mal, maleyl; Bz, benzoyl; OMe, methyl ester; OEt, ethyl ester) and a series of amino-acid amides. Most of the reactions studied can be described by the simplest kinetic model without the nucleophile binding to the acyl-enzyme. The alpha-chymotrypsin-catalyzed transfer of the Mal-L-Ala-L-Ala-L-Phe group to the amides of L-Phe and L-Tyr showed a linear dependence of the partition constant, p, on the nucleophile concentration which can be interpreted by the hydrolysis of the acyl-enzyme-nucleophile complex.

[Metalloproteinase of Bacillus mesentericus, strain V-313].

A homogeneous metalloproteinase has been isolated with a 28% yield from the culture fluid of Bacillus mesentericus, strain B-313. The isolation procedure included chromatography on bacitracin-silochrome and gel filtration on Acrylex P-10 and Sephadex G-75. The enzyme has a molecular mass of 41,000 Da; its N-terminal sequence, which appears as A-A-T-T-G-T-G-T-T-L-K-G-K-T-V-S-L-N-I, is identical with that of the B.

[Isolation and characteristics of Bacillus megaterium metalloproteinase].

Stepwise application of affinity chromatography on bacitracin-silochrome, gel filtration on Acrylex P-10, rechromatography on bacitracin-Sepharose 4B and gel filtration on Sephadex G-15, a homogeneous metalloproteinase (M(r) = 35,000 Da) has been isolated from the cultural filtrate of B. megaterium strain 599. The amino acid composition and N-terminal sequence (20 amino acids) of the enzyme have been determined.

How to predict product yield in kinetically controlled enzymatic peptide synthesis.

The published theory of kinetically controlled enzymatic peptide synthesis needed experimental verification. We carried out the measurement of the peptide yield and estimation of the key parameters alpha and beta for papain-catalyzed synthesis of Mal-L-Phe-L-Ala-LLeuNH(2) from Mal-L-Phe-L-AlaOMe and L-LeuNH(2). The experimental results demonstrate that this theory adequately describes the real process.