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Journal article
The Primitive Protozoon Trichomonas vaginalisContains Two Methionine γ-Lyase Genes That Encode Members of the γ-Family of Pyridoxal 5′-Phosphate-dependent Enzymes
The Journal of biological chemistry, v 273(10), pp 5549-5556
06 Mar 1998
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Abstract
Methionine γ-lyase, the enzyme that catalyzes the breakdown of methionine by an α,γ-elimination reaction and is a member of the γ-family of pyridoxal 5′-phosphate-dependent enzymes, is present in high activity in the primitive protozoan parasite Trichomonas vaginalisbut is absent from mammals. Two genes, mgl1 and mgl2, encoding methionine γ-lyase, have now been isolated from T. vaginalis. They are both single copy, encode predicted proteins (MGL1 and MGL2) of 43 kDa, have 69% sequence identity with each other, and show a high degree of sequence identity to methionine γ-lyase from Pseudomonas putida (44%) and other related pyridoxal 5′-phosphate-dependent enzymes such as human cystathionine γ-lyase (42%) and Escherichia coli cystathionine β-lyase (30%). mgl1 and mgl2 have been expressed in E. coli as a fusion with a six-histidine tag and the recombinant proteins (rMGL1 and rMGL2) purified by metal-chelate affinity chromatography. rMGL1 and rMGL2 were found to have high activity toward methionine (10.4 and 0.67 μmol/min/mg of protein, respectively), homocysteine (370 and 128 μmol/min/mg of protein), cysteine (6.02 and 1.06 μmol/min/mg of protein), and O-acetylserine (3.74 and 1.51 μmol/min/mg of protein), but to be inactive toward cystathionine. Site-directed mutagenesis of an active site cysteine (C113G for MGL1 and C116G for MGL2) reduced the activity of the recombinant enzymes toward both methionine and homocysteine by approximately 80% (rMGL1) and 90% (rMGL2). In contrast, the activity of mutated rMGL2 toward cysteine and O-acetylserine was increased (to 214 and 142%, respectively), whereas that of mutated rMGL1 was reduced to 39 and 49%, respectively. These findings demonstrate the importance of this cysteine residue in the α,β-elimination and α,γ-elimination reactions catalyzed by trichomonad methionine γ-lyase.
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- Title
- The Primitive Protozoon Trichomonas vaginalisContains Two Methionine γ-Lyase Genes That Encode Members of the γ-Family of Pyridoxal 5′-Phosphate-dependent Enzymes
- Creators
- Amanda E. McKie - University of GlasgowThomas Edlind - Allegheny University of the Health SciencesJohn Walker - Wellcome Unit of Molecular Parasitology, University of Glasgow, The Anderson College, Glasgow G11 6NU, Scotland, United KingdomJeremy C. Mottram - Wellcome Unit of Molecular Parasitology, University of Glasgow, The Anderson College, Glasgow G11 6NU, Scotland, United Kingdom.Graham H. Coombs - University of GlasgowJoseph D Walker - Thomas R. Kline School of Law (2014-)
- Publication Details
- The Journal of biological chemistry, v 273(10), pp 5549-5556
- Publisher
- Elsevier
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Thomas R. Kline School of Law
- Web of Science ID
- WOS:000072345000023
- Scopus ID
- 2-s2.0-0032489551
- Other Identifier
- 991019168613404721
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- Collaboration types
- Domestic collaboration
- International collaboration
- Web of Science research areas
- Biochemistry & Molecular Biology