Abstract
Pyrroline-5-carboxylate reductase (P5CR) plays an important role in the survival of Mycobacterium tuberculosis and is related to virulence of this pathogen. RT-PCR analysis indicated that proC, encoding P5CR, was expressed at the transcriptional level cultured in vitro. The His-rMtP5CR with an N-terminal His-tag (His-rMtP5CR) was firstly purified in Escherichia coli and rMtP5CR was obtained by removal of the N-terminal fusion partner using enterokinase. His-rMtP5CR had considerable beta-pleated sheet analyzed by circular dichroism spectroscopy. The effect of pH, temperature, cations, denaturants, and detergents on the purified enzyme activity and stability was characterized. The N-terminal fusion partner was found to have very little effect on the biochemical properties of P5CR.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Cells, Cultured
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Circular Dichroism
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Cloning, Molecular
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DNA / genetics
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Escherichia coli / cytology
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Escherichia coli / genetics
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Escherichia coli / metabolism
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Gene Expression Regulation, Enzymologic
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Hydrogen-Ion Concentration
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Kinetics
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Mycobacterium tuberculosis / enzymology*
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Pyrroline Carboxylate Reductases / chemistry*
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Pyrroline Carboxylate Reductases / genetics
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Pyrroline Carboxylate Reductases / isolation & purification
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RNA / genetics
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / isolation & purification
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Reverse Transcriptase Polymerase Chain Reaction / methods
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Temperature
Substances
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Recombinant Fusion Proteins
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RNA
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DNA
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Pyrroline Carboxylate Reductases
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pyrroline-5-carboxylate reductase, M tuberculosis