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Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis

BACKGROUND: Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorg...

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Autores principales: Venos, Erik S, Knodel, Marvin H, Radford, Cynthia L, Berger, Bradley J
Formato: Texto
Lenguaje:English
Publicado: BioMed Central 2004
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC524486/
https://www.ncbi.nlm.nih.gov/pubmed/15471546
http://dx.doi.org/10.1186/1471-2180-4-39
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author Venos, Erik S
Knodel, Marvin H
Radford, Cynthia L
Berger, Bradley J
author_facet Venos, Erik S
Knodel, Marvin H
Radford, Cynthia L
Berger, Bradley J
author_sort Venos, Erik S
collection PubMed
description BACKGROUND: Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorganisms. The final step of this pathway, the conversion of ketomethiobutyrate to methionine, can be performed by aspartate, tyrosine, or branched-chain amino acid aminotransferases depending on the particular species examined. RESULTS: The gene encoding for branched-chain amino acid aminotransferase in Mycobacterium tuberculosis H37Rv has been cloned, expressed, and characterised. The enzyme was found to be a member of the aminotransferase IIIa subfamily, and closely related to the corresponding aminotransferase in Bacillus subtilis, but not to that found in B. anthracis or B. cereus. The amino donor preference for the formation of methionine from ketomethiobutyrate was for isoleucine, leucine, valine, glutamate, and phenylalanine. The enzyme catalysed branched-chain amino acid and ketomethiobutyrate transamination with a Km of 1.77 – 7.44 mM and a Vmax of 2.17 – 5.70 μmol/min/mg protein, and transamination of ketoglutarate with a Km of 5.79 – 6.95 mM and a Vmax of 11.82 – 14.35 μmol/min/mg protein. Aminooxy compounds were examined as potential enzyme inhibitors, with O-benzylhydroxylamine, O-t-butylhydroxylamine, carboxymethoxylamine, and O-allylhydroxylamine yielding mixed-type inhibition with Ki values of 8.20 – 21.61 μM. These same compounds were examined as antimycobacterial agents against M. tuberculosis and a lower biohazard M. marinum model system, and were found to completely prevent cell growth. O-Allylhydroxylamine was the most effective growth inhibitor with an MIC of 78 μM against M. marinum and one of 156 μM against M. tuberculosis. CONCLUSION: Methionine formation from ketomethiobutyrate is catalysed by a branched-chain amino acid aminotransferase in M. tuberculosis. This enzyme can be inhibited by selected aminooxy compounds, which also have effectiveness in preventing cell growth in culture. These compounds represent a starting point for the synthesis of branched-chain aminotransferase inhibitors with higher activity and lower toxicity.
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spelling pubmed-5244862004-10-31 Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis Venos, Erik S Knodel, Marvin H Radford, Cynthia L Berger, Bradley J BMC Microbiol Research Article BACKGROUND: Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorganisms. The final step of this pathway, the conversion of ketomethiobutyrate to methionine, can be performed by aspartate, tyrosine, or branched-chain amino acid aminotransferases depending on the particular species examined. RESULTS: The gene encoding for branched-chain amino acid aminotransferase in Mycobacterium tuberculosis H37Rv has been cloned, expressed, and characterised. The enzyme was found to be a member of the aminotransferase IIIa subfamily, and closely related to the corresponding aminotransferase in Bacillus subtilis, but not to that found in B. anthracis or B. cereus. The amino donor preference for the formation of methionine from ketomethiobutyrate was for isoleucine, leucine, valine, glutamate, and phenylalanine. The enzyme catalysed branched-chain amino acid and ketomethiobutyrate transamination with a Km of 1.77 – 7.44 mM and a Vmax of 2.17 – 5.70 μmol/min/mg protein, and transamination of ketoglutarate with a Km of 5.79 – 6.95 mM and a Vmax of 11.82 – 14.35 μmol/min/mg protein. Aminooxy compounds were examined as potential enzyme inhibitors, with O-benzylhydroxylamine, O-t-butylhydroxylamine, carboxymethoxylamine, and O-allylhydroxylamine yielding mixed-type inhibition with Ki values of 8.20 – 21.61 μM. These same compounds were examined as antimycobacterial agents against M. tuberculosis and a lower biohazard M. marinum model system, and were found to completely prevent cell growth. O-Allylhydroxylamine was the most effective growth inhibitor with an MIC of 78 μM against M. marinum and one of 156 μM against M. tuberculosis. CONCLUSION: Methionine formation from ketomethiobutyrate is catalysed by a branched-chain amino acid aminotransferase in M. tuberculosis. This enzyme can be inhibited by selected aminooxy compounds, which also have effectiveness in preventing cell growth in culture. These compounds represent a starting point for the synthesis of branched-chain aminotransferase inhibitors with higher activity and lower toxicity. BioMed Central 2004-10-07 /pmc/articles/PMC524486/ /pubmed/15471546 http://dx.doi.org/10.1186/1471-2180-4-39 Text en Copyright © 2004 Venos et al; licensee BioMed Central Ltd.
spellingShingle Research Article
Venos, Erik S
Knodel, Marvin H
Radford, Cynthia L
Berger, Bradley J
Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title_full Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title_fullStr Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title_full_unstemmed Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title_short Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis
title_sort branched-chain amino acid aminotransferase and methionine formation in mycobacterium tuberculosis
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC524486/
https://www.ncbi.nlm.nih.gov/pubmed/15471546
http://dx.doi.org/10.1186/1471-2180-4-39
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