Biomedical subjects

M Prucha

Publications and source records attributed to M Prucha.

10 recordsLinked to original sources

PubMed1997

Evidence for an isomeric muconolactone isomerase involved in the metabolism of 4-methylmuconolactone by Alcaligenes eutrophus JMP134.

An enzyme specifically induced during 4-methylmuconolactone metabolism by Alcaligenes eutrophus JMP 134 and that exhibited muconolactone isomerizing activity was purified to homogeneity. The enzyme, involved in the isomerization of 3-methylmuconolactone had a high degree of sequence similarity with muconolactone isomerase of Alcaligenes eutrophus JMP 134 and other previously described muconolactone isomerases of the 3-oxoadipate pathway. Kinetic analysis showed that the enzyme has a substrate spectrum and a reaction mechanism similar to those of the muconolactone isomerase, but that it has distinct kinetic properties.

Alcaligenes↗

PubMed1996

Muconolactone isomerase of the 3-oxoadipate pathway catalyzes dechlorination of 5-chloro-substituted muconolactones.

An enzyme of Alcaligenes eutrophus JMP 134 which catalyzes dechlorination of (4R, 5R)- and (4R,5S)-5-chloro-3-methyl- and (4R, 5S)-5-chloromuconolactone of principally 3-methyl-trans-, 3-methyl-cis-dienelactone and cis-dienelactone, respectively, was purified to homogeneity. The enzyme was identified as muconolactone isomerase on the basis of its high activity with muconolactone and on its high degree of sequence similarity with previously described muconolactone isomerases. Molecular mass determinations of the highly hydrophobic and heat-resistant enzyme indicated a decameric structure involving a single 10.100-kDa subunit similar to that of muconolactone isomerase of Pseudomonas putida. Kinetic analysis showed cooperative effects between the subunits during conversion of (4R, 5S)-5-chloro-3 -methylmuconolactone. (4R, 5S)-5-chloromuconolactone was the preferred substrate, over the natural substrate (4S)-muconolactone. The (4S, 5S)-structure was found to be an inhibitor of (4R, 5R)-5-chloro-3-methylmuconolactone transformation. Methylsubstitution of the substrate results in a higher affinity for the enzyme, but a drastically lower velocity, resulting in a lower specificity constant.

4-Butyrolactone↗

PubMed1996

Metabolism of 5-chlorosubstituted muconolactones.

The stereochemistry of the four stereoforms of 5-chloro-3-methylmuconolactones could be deduced from NMR and stability data, and from the comparison with authentic (4R, 5S)-5-chloromuconolactone. Muconolactone isomerase of Alcaligenes eutrophus JMP 134 was shown to catalyze syn-elimination of hydrogen chloride from (4R, 5R)-5-chloro-3-methylmuconolactone, (4R, 5S)-5-chloro-3 -methylmuconolactone and (4R, 5S)-5-chloromuconolactone to form 3-methyl-trans-dienelactone, 3-methyl-cis-dienelactone and a 3:1 mixture of cis- and trans-dienelactone, respectively. 3-Methyl-trans-dienelactone was a substrate of pJP4-encoded dienelactone hydrolase of A. eutrophus JMP 134, whereas 3-methyl-cis-dienelactone transformation was negligible indicating a restricted substrate specificity of this enzyme. Both substrates were transformed into 3-methylmaleylacetate which in turn was a substrate for maleylacetate reductase. This compound was shown to possess a cyclic structure (4-hydroxy-3-methyl-muconolactone) under acidic conditions.

4-Butyrolactone↗

PubMed1994

Analysis of three 2,3-dihydroxybiphenyl 1,2-dioxygenases found in Rhodococcus globerulus P6. Identification of a new family of extradiol dioxygenases.

The polychlorobiphenyl-degrading bacterium Rhodococcus globerulus P6 contains three bphC genes encoding 2,3-dihydroxybiphenyl 1,2-dioxygenases. One of them, bphC1, is clustered with the bphB gene which encodes 2,3-dihydroxy-4-phenylhexa-4,6-diene dehydrogenase and constitutes part of the bph operon specifying the degradation of biphenyl. The nucleotide sequence of bphB and the three bphC genes has been determined. The protein products of the bphBC1 gene cluster were found to exhibit significant homology with the corresponding proteins of analogous degradative pathways in Gram-negative bacteria; the highest homology was in those of the toluene degradation pathway of Pseudomonas putida strain F1. No homology was found between bphC2 and bphC3 and any other sequence in the database. At least two of the three meta cleavage enzymes are inducible by biphenyl. 2,3-Dihydroxybiphenyl 1,2-dioxygenase II, encoded by the bphC2 gene, was purified to apparent homogeneity from a recombinant Escherichia coli strain. The enzyme differed from other extradiol dioxygenases in having a subunit molecular mass of 21 kDa and a hexameric structure. The enzyme contains one tightly bound iron per subunit. These characteristics demonstrate that the 2,3-dihydroxybiphenyl 1,2-dioxygenases encoded by bphC2 and bphC3 belong to a new class of extradiol dioxygenases.

Amino Acid Sequence↗

PubMed1986