Publications by authors named "N A Stambolieva"
The Zn-proteinase, isolated from Saccharomonospora canescens (NPS), shares many common features with thermolysin, but considerable differences are also evident, as far as the substrate recognition site is concerned. In substrates of general structure AcylAlaAlaPhe 4NA, this neutral proteinase cleaves only the arylamide bond (non-typical activity of Zn-proteinases), while thermolysin attacks the peptide bond Ala-Phe. Phosphoramidon is a powerful tight binding inhibitor for thermolysin and significantly less specific towards NPS.
View Article and Find Full Text PDFA new set of experimental kinetic data on the hydrolysis of a series of phenylacetyl p-substituted anilides catalyzed by penicillin G acylase from Escherichia coli (PGA) is presented in this article. The Hammett plot of log(k(cat,R)/k(cat,H)) versus sigma(p) (-) has three linear segments, which distinguishes the enzyme from the other N-terminal nucleophile hydrolases for which data are available. Three amino acids in the vicinity of the catalytic SerB1 (AsnB241, AlaB69, and GlnB23) were included in the quantum mechanical model.
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- Kinetic studies show that a polar group in the amine part of phenylacetyl-arylamides is essential for their correct positioning in the penicillin acylase enzyme of E. coli.
- Quantum mechanical modeling reveals that substrates with asymmetry bind in two different, non-symmetrical ways to the positively charged arginine residues.
- This suggests that the arginine pair (ArgA145 and ArgB263) plays a key role in determining how substrates are selected and oriented by penicillin acylase.
Article Synopsis
- The study investigated the arylamidase activity of Zn-proteinase from Saccharomonospora canescens (NPS) using peptide nitroanilides with different amino acid sequences and N-acyl groups.
- Key findings included that variations in substrate structure impact catalytic efficiency, particularly highlighting the significance of interactions between specific substrate positions (P(1) and P(4)) and corresponding enzyme subsites (S(1) and S(4)).
- The arylamidase activity in Zn-proteinases correlates with their active site structure, which features penta-coordinated Zn(2+) and a tyrosyl residue that helps bind substrates and stabilize reaction intermediates.
The methyl ester of 2-benzoxazolon-3-yl-acetic acid was used as an acyl donor in the penicillin amidase-catalysed transfer reaction to 7-aminodesacetoxycephalosporanic acid. The synthesis of 7-(2-benzoxazolon-3-yl-acetamido)-desacetoxycephalosporanic acid was carried out as a kinetically controlled reaction. A characteristic feature of this system is that the benzoxazolone derivatives are very low specific substrates for penicillin amidase (the kcat/Km values for their hydrolysis were shown to be 10(5)-fold lower compared to the corresponding values for phenylacetyl derivatives).
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