PMID:21696459
| Citation |
Subedi, KP, Choi, D, Kim, I, Min, B and Park, C (2011) Hsp31 of Escherichia coli K-12 is glyoxalase III. Mol. Microbiol. 81:926-36 |
|---|---|
| Abstract |
Hsp31 encoded by hchA is known as a heat-inducible molecular chaperone. Although structure studies revealed that Hsp31 has a putative catalytic triad consisting of Asp-214, His-186 and Cys-185, its enzymatic function, besides weak amino-peptidase activity, is still unknown. We found that Hsp31 displays glyoxalase activity that catalyses the conversion of methylglyoxal (MG) to d-lactate without an additional cofactor. The glyoxalase activity was completely abolished in the hchA-deficient strain, confirming the relationship between the hchA gene and its enzymatic activity in vivo. Hsp31 exhibits Michaelis-Menten kinetics for substrates MG with K(m) and k(cat) of 1.43±0.12 mM and 156.9±5.5 min⁻¹ respectively. The highest glyoxalase activity was found at 35-40 °C and pH of 6.0-8.0, and the activity was significantly inhibited by Cu²⁺, Fe³⁺ and Zn²⁺. Mutagenesis studies based on our evaluation of conserved catalytic residues revealed that the Cys-185 and Glu-77 were essential for catalysis, whereas His-186 was less crucial for enzymatic function, although it participates in the catalytic process. The stationary-phase Escherichia coli cells became more susceptible to MG when hchA was deleted, which was complemented by an expression of plasmid-encoded hchA. Furthermore, an accumulation of intracellular MG was observed in hchA-deficient strains. |
| Links |
PubMed Online version:10.1111/j.1365-2958.2011.07736.x |
| Keywords |
Aldehyde Oxidoreductases/genetics; Aldehyde Oxidoreductases/metabolism; Amino Acid Substitution; Biotransformation; Cations, Divalent/metabolism; DNA Mutational Analysis; Enzyme Inhibitors/metabolism; Escherichia coli K12/enzymology; Escherichia coli K12/genetics; Escherichia coli Proteins/genetics; Escherichia coli Proteins/metabolism; Gene Deletion; Genetic Complementation Test; Hydrogen-Ion Concentration; Kinetics; Lactates/metabolism; Metals/metabolism; Models, Molecular; Molecular Chaperones/genetics; Molecular Chaperones/metabolism; Mutagenesis, Site-Directed; Pyruvaldehyde/metabolism; Temperature |
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