Identification and characterization of bifunctional proline racemase/hydroxyproline epimerase from archaea: discrimination of substrates and molecular evolution

PloS One
Seiya WatanabeYasuo Watanabe

Abstract

Proline racemase (ProR) is a member of the pyridoxal 5'-phosphate-independent racemase family, and is involved in the Stickland reaction (fermentation) in certain clostridia as well as the mechanisms underlying the escape of parasites from host immunity in eukaryotic Trypanosoma. Hydroxyproline epimerase (HypE), which is in the same protein family as ProR, catalyzes the first step of the trans-4-hydroxy-L-proline metabolism of bacteria. Their substrate specificities were previously considered to be very strict, in spite of similarities in their structures and catalytic mechanisms, and no racemase/epimerase from the ProR superfamily has been found in archaea. We here characterized the ProR-like protein (OCC_00372) from the hyperthermophilic archaeon, Thermococcus litoralis (TlProR). This protein could reversibly catalyze not only the racemization of proline, but also the epimerization of 4-hydroxyproline and 3-hydroxyproline with similar kinetic constants. Among the four (putative) ligand binding sites, one amino acid substitution was detected between TlProR (tryptophan at the position of 241) and natural ProR (phenylalanine). The W241F mutant showed a significant preference for proline over hydroxyproline, suggesting that this ...Continue Reading

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Citations

Feb 14, 2015·Journal of Biotechnology·Seiya WatanabeYasuo Watanabe
Dec 9, 2016·Scientific Reports·Seiya WatanabeYasuo Watanabe
Feb 3, 2016·Journal of Bacteriology·Siyun ChenTurlough M Finan
Oct 1, 2015·Environmental Microbiology·Sara B Hernández, Felipe Cava
Apr 16, 2020·Applied Microbiology and Biotechnology·Ryotaro Hara, Kuniki Kino
Aug 4, 2021·Amino Acids·Shengdi HuGuoyao Wu

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Methods Mentioned

BETA
PCR
gel filtration

Software Mentioned

Clustal W
ImageJ
BLAST
- BLAST
Protein

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