  Delta 2- and delta 3-cephalosporins, penicillinate and 6-unsubstituted penems. Intrinsic reactivity and interaction with beta-lactamases and D-alanyl-D-alanine-cleaving serine peptidasesFrère, Jean-Marie  ; ; et alin Biochemical Journal (1982), 203(1), 223-234 The intrinsic reactivity of delta 2- and delta 3-deacetoxy-7-phenylacetamidocephalosporanates, penicillanate, unsubstituted, 2-methyl- and 2-phenyl-penems and other beta-lactam antibiotics has been ... [more ▼] The intrinsic reactivity of delta 2- and delta 3-deacetoxy-7-phenylacetamidocephalosporanates, penicillanate, unsubstituted, 2-methyl- and 2-phenyl-penems and other beta-lactam antibiotics has been expressed in terms of the second-order rate constant (M-1.s-1(OH-)) for the hydrolysis of the beta-lactam amide bond by OH- at 37 degrees C. The values thus obtained have been compared with the second-order rate constants (M-1.s-1(enzyme) for the opening of the same beta-lactam amide bond during interaction with the beta-lactamases of Streptomyces albus G and Actinomadura R39 and the D-alanyl-D-alanine-cleaving serine peptidases of Streptomyces R61 and Actinomadura R39. Depending on the cases, the accelerating effect due to enzyme action and expressed by the ratio M-1.s-1(enzyme)/M-1.s-1(OH) varies from less than 2 to more than 1 x 10(6). The primary parameter that governs enzyme action is the goodness of fit of the beta-lactam molecule to the enzyme cavity rather than its intrinsic reactivity. With the D-alanyl-D-alanine-cleaving serine peptidases, the three penems studied form intermediate complexes characterized by very short half lives of 14-100 s, values significantly lower than those exhibited by most beta-lactam compounds. [less ▲] Detailed reference viewed: 7 (0 ULg)Stability of D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid in relation to its possible occurrence as a degradation product of penicillin by the exocellular DD-carboxypeptidase-transpeptidase from Streptomyces R61 and the membrane-bound dd-carboxypeptidase from Bacillus stearothermophilus; ; Frère, Jean-Marie et alin Journal of Biological Chemistry (1978), 253(10), 3660-3665 The stability of D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid has been studied under various conditions. In 10 mM cacodylate, pH 6.5, and at 55 degrees C, D-5,5-dimethyl-delta2-thiazoline-4 ... [more ▼] The stability of D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid has been studied under various conditions. In 10 mM cacodylate, pH 6.5, and at 55 degrees C, D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid (at concentrations lower than 1 mM) is hydrolyzed into N-formyl-D-penicillamine with a half-life of 3 to 4 min. On this basis, it is very unlikely that D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid could be one of the end products resulting from the cleavage of benzylpenicillin by the DD-carboxypeptidase of Bacillus stearothermophilus (as reported by Hammarstrom and Strominger (1976) J. Biol. Chem. 251, 7947--7949). In 3 mM phosphate, pH 7.5, and at 37 degrees C, D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid (at concentrations lower than 1 mM) has a half-life of 45 min. On the basis of kinetic experiments carried out under these conditions with phenoxymethylpenicillin and the DD-carboxypeptidase-transpeptidase of Streptomyces R61, it is concluded that the primary product which arises from the thiazolidine moiety of the antibiotic molecule and gives rise to N-formyl-D-penicillamine, has a half-life of 10 min, a value which is not compatible with the hypothesis that D-5,5-dimethyl-delta2-thiazoline-4-carboxylic acid would be an intermediate involved in the fragmentation pathway. [less ▲] Detailed reference viewed: 5 (0 ULg) Interactions between beta-lactam antibiotics and isolated membranes of Streptococcus faecalis ATCC 9790; Ghuysen, Jean-Marie ; Binot, Françoise et alin European Journal of Biochemistry (1977), 75(1), 231-239 The DD-carboxypeptidase-exchange membrane-bound enzyme in Streptococcus faecalis ATCC 9790 reacts with beta-lactam antibiotics to form complexes with rather long half-lives. Depending upon the antibiotic ... [more ▼] The DD-carboxypeptidase-exchange membrane-bound enzyme in Streptococcus faecalis ATCC 9790 reacts with beta-lactam antibiotics to form complexes with rather long half-lives. Depending upon the antibiotic, the second-order rate constants for complex formation range from 0.75-560 M-1 S-1 (at 37 degrees C and in water) and the first-order rate constants for complex breakdown range from 1.3 to 26 x 10(-5) s-1 (at 37 degrees C and in 5 mM phosphate buffer pH 7.5). There are about 30 pmol of DD-carboxypeptidase-exchange enzyme per mg of membrane protein. The degradation products arising from benzylpenicillin are phenylacetylglycine and probably N-formyl-D-penicillamine. Isolated membranes also contain other penicillin binding sites (about 70 pmol/mg membrane protein). That part of benzylpenicillin which reacts with at least some of these latter sites is slowly degraded into penicilloic acid. Normal functioning of the DD-carboxypeptidase-exchange membrane-bound enzyme is important, if not essential, for cell growth. With the beta-lactam antibiotics tested inhibition of cell growth is mainly related to the rates of formation of the inactive enzyme-antibiotic complexes. The relationship, however, is not a direct one probably due to the competitive effect exerted by the other penicillin binding sites. [less ▲] Detailed reference viewed: 4 (0 ULg)Fate of thiazolidine ring during fragmentation of penicillin by exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61Frère, Jean-Marie ; Ghuysen, Jean-Marie ; et alin Nature (1976), 260(5550), 451-454 LIKE various beta-lactamases, acylases and esterases1, the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61 degrades benzylpenicillin and other beta-lactam antibiotics2-4. The R61 enzyme ... [more ▼] LIKE various beta-lactamases, acylases and esterases1, the exocellular DD-carboxypeptidase-transpeptidase of Streptomyces R61 degrades benzylpenicillin and other beta-lactam antibiotics2-4. The R61 enzyme, however, markedly differs from the other penicillin-degrading enzymes in causing fragmentation of the penicillin nucleus. By using 8-14C-benzylpenicillin (benzyl labelled) as substrate, one of the fragments produced was shown to be 14C-phenylacetylglycine5. The reaction with the R61 enzyme is also peculiar in that it is a slow process. This is because of the long half life of the stoichiometnc complex transitorily formed between the antibiotic and the enzyme. Thus, for example, the value of the half life for the complex formed with benzylpenicillin is 80 min in 10 mM phosphate buffer (pH 7.0) and at 37 °C. As breakdown of the complex proceeds, however, phenylacetylglycine (when benzylpenicillin is used as substrate) is released and the enzyme concomitantly recovers its ability to bind penicillin. We have now characterised the fragment (hereby designated as the Y product) arising from the thiazolidine ring of penicillin as a result of the fragmentation of the antibiotic molecule by the R61 enzyme. [less ▲] Detailed reference viewed: 11 (1 ULg) |
||
