Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it>
<p>Abstract</p> <p>Background</p> <p>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 biosyn...
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| Format: | Article |
| Language: | English |
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BMC
2004-10-01
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| Series: | BMC Microbiology |
| Online Access: | http://www.biomedcentral.com/1471-2180/4/39 |
| _version_ | 1818663139848224768 |
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| author | Radford Cynthia L Knodel Marvin H Venos Erik S Berger Bradley J |
| author_facet | Radford Cynthia L Knodel Marvin H Venos Erik S Berger Bradley J |
| author_sort | Radford Cynthia L |
| collection | DOAJ |
| description | <p>Abstract</p> <p>Background</p> <p>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.</p> <p>Results</p> <p>The gene encoding for branched-chain amino acid aminotransferase in <it>Mycobacterium tuberculosis </it>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 <it>Bacillus subtilis</it>, but not to that found in <it>B. anthracis </it>or <it>B. cereus</it>. 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 <it>M. tuberculosis </it>and a lower biohazard <it>M. marinum </it>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 <it>M. marinum </it>and one of 156 ��M against <it>M. tuberculosis</it>.</p> <p>Conclusion</p> <p>Methionine formation from ketomethiobutyrate is catalysed by a branched-chain amino acid aminotransferase in <it>M. tuberculosis</it>. 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.</p> |
| first_indexed | 2024-12-17T05:12:06Z |
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| id | doaj.art-ea099eb0f67d432bbc049493ba973467 |
| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-12-17T05:12:06Z |
| publishDate | 2004-10-01 |
| publisher | BMC |
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| series | BMC Microbiology |
| spelling | doaj.art-ea099eb0f67d432bbc049493ba9734672022-12-21T22:02:15ZengBMCBMC Microbiology1471-21802004-10-01413910.1186/1471-2180-4-39Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it>Radford Cynthia LKnodel Marvin HVenos Erik SBerger Bradley J<p>Abstract</p> <p>Background</p> <p>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.</p> <p>Results</p> <p>The gene encoding for branched-chain amino acid aminotransferase in <it>Mycobacterium tuberculosis </it>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 <it>Bacillus subtilis</it>, but not to that found in <it>B. anthracis </it>or <it>B. cereus</it>. 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 <it>M. tuberculosis </it>and a lower biohazard <it>M. marinum </it>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 <it>M. marinum </it>and one of 156 ��M against <it>M. tuberculosis</it>.</p> <p>Conclusion</p> <p>Methionine formation from ketomethiobutyrate is catalysed by a branched-chain amino acid aminotransferase in <it>M. tuberculosis</it>. 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.</p>http://www.biomedcentral.com/1471-2180/4/39 |
| spellingShingle | Radford Cynthia L Knodel Marvin H Venos Erik S Berger Bradley J Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> BMC Microbiology |
| title | Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> |
| title_full | Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> |
| title_fullStr | Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> |
| title_full_unstemmed | Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> |
| title_short | Branched-chain amino acid aminotransferase and methionine formation in <it>Mycobacterium tuberculosis</it> |
| title_sort | branched chain amino acid aminotransferase and methionine formation in it mycobacterium tuberculosis it |
| url | http://www.biomedcentral.com/1471-2180/4/39 |
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