Characterization of a novel D-lyxose isomerase from Cohnella laevoribosii RI-39 sp. nov.

Eun Ah Cho, Dong Woo Lee, Yun Hwan Cha, Sang Jae Lee, Heung Chae Jung, Jae Gu Pan, Yu Ryang Pyun

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

A newly isolated bacterium, Cohnella laevoribosii RI-39, could grow in a defined medium with L-ribose as the sole carbon source. A 21-kDa protein isomerizing L-ribose to L-ribulose, as well as D-lyxose to D-xylulose, was purified to homogeneity from this bacterium. Based on the N-terminal and internal amino acid sequences of the purified enzyme obtained by N-terminal sequencing and quantitative time of flight mass spectrometry-mass spectrometry analyses, a 549-bp gene (lyxA) encoding D-lyxose (L-ribose) isomerase was cloned and expressed in Escherichia coli. The purified endogenous enzyme and the recombinant enzyme formed homodimers that were activated by Mn2+. C. laevoribosii D-lyxose (L-ribose) isomerase (CLLI) exhibits maximal activity at pH 6.5 and 70°C in the presence of Mn2+ for D-lyxose and L-ribose, and its isoelectric point (pI) is 4.2 (calculated pI, 4.9). The enzyme is specific for D-lyxose, L-ribose, and D-mannose, with apparent Km values of 22.4 ± 1.5 mM, 121.7 ± 10.8 mM, and 34.0 ± 1.1 mM, respectively. The catalytic efficiencies (kcat/Km) of CLLI were 84.9 ± 5.8 mM-1 s-1 for D-lyxose (V max, 5,434.8 U mg-1), 0.2 mM-1 s-1 for L-ribose (Vmax, 75.5 ± 6.0 U mg-1), and 1.4 ± 0.1 mM-1 s-1 for D-mannose (Vmax, 131.8 ± 7.4 U mg-1). The ability of lyxA to permit E. coli cells to grow on D-lyxose and L-ribose and homology searches of other sugar-related enzymes, as well as previously described sugar isomerases, suggest that CLLI is a novel type of rare sugar isomerase.

Original languageEnglish
Pages (from-to)1655-1663
Number of pages9
JournalJournal of Bacteriology
Volume189
Issue number5
DOIs
Publication statusPublished - 2007 Mar 1

Fingerprint

Isomerases
Ribose
Enzymes
Mannose
Xylulose
Escherichia coli
Bacteria
Isoelectric Point
Tandem Mass Spectrometry
lyxose
Amino Acid Sequence
Carbon
Genes

All Science Journal Classification (ASJC) codes

  • Microbiology
  • Molecular Biology

Cite this

Cho, Eun Ah ; Lee, Dong Woo ; Cha, Yun Hwan ; Lee, Sang Jae ; Jung, Heung Chae ; Pan, Jae Gu ; Pyun, Yu Ryang. / Characterization of a novel D-lyxose isomerase from Cohnella laevoribosii RI-39 sp. nov. In: Journal of Bacteriology. 2007 ; Vol. 189, No. 5. pp. 1655-1663.
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abstract = "A newly isolated bacterium, Cohnella laevoribosii RI-39, could grow in a defined medium with L-ribose as the sole carbon source. A 21-kDa protein isomerizing L-ribose to L-ribulose, as well as D-lyxose to D-xylulose, was purified to homogeneity from this bacterium. Based on the N-terminal and internal amino acid sequences of the purified enzyme obtained by N-terminal sequencing and quantitative time of flight mass spectrometry-mass spectrometry analyses, a 549-bp gene (lyxA) encoding D-lyxose (L-ribose) isomerase was cloned and expressed in Escherichia coli. The purified endogenous enzyme and the recombinant enzyme formed homodimers that were activated by Mn2+. C. laevoribosii D-lyxose (L-ribose) isomerase (CLLI) exhibits maximal activity at pH 6.5 and 70°C in the presence of Mn2+ for D-lyxose and L-ribose, and its isoelectric point (pI) is 4.2 (calculated pI, 4.9). The enzyme is specific for D-lyxose, L-ribose, and D-mannose, with apparent Km values of 22.4 ± 1.5 mM, 121.7 ± 10.8 mM, and 34.0 ± 1.1 mM, respectively. The catalytic efficiencies (kcat/Km) of CLLI were 84.9 ± 5.8 mM-1 s-1 for D-lyxose (V max, 5,434.8 U mg-1), 0.2 mM-1 s-1 for L-ribose (Vmax, 75.5 ± 6.0 U mg-1), and 1.4 ± 0.1 mM-1 s-1 for D-mannose (Vmax, 131.8 ± 7.4 U mg-1). The ability of lyxA to permit E. coli cells to grow on D-lyxose and L-ribose and homology searches of other sugar-related enzymes, as well as previously described sugar isomerases, suggest that CLLI is a novel type of rare sugar isomerase.",
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Characterization of a novel D-lyxose isomerase from Cohnella laevoribosii RI-39 sp. nov. / Cho, Eun Ah; Lee, Dong Woo; Cha, Yun Hwan; Lee, Sang Jae; Jung, Heung Chae; Pan, Jae Gu; Pyun, Yu Ryang.

In: Journal of Bacteriology, Vol. 189, No. 5, 01.03.2007, p. 1655-1663.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Characterization of a novel D-lyxose isomerase from Cohnella laevoribosii RI-39 sp. nov.

AU - Cho, Eun Ah

AU - Lee, Dong Woo

AU - Cha, Yun Hwan

AU - Lee, Sang Jae

AU - Jung, Heung Chae

AU - Pan, Jae Gu

AU - Pyun, Yu Ryang

PY - 2007/3/1

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N2 - A newly isolated bacterium, Cohnella laevoribosii RI-39, could grow in a defined medium with L-ribose as the sole carbon source. A 21-kDa protein isomerizing L-ribose to L-ribulose, as well as D-lyxose to D-xylulose, was purified to homogeneity from this bacterium. Based on the N-terminal and internal amino acid sequences of the purified enzyme obtained by N-terminal sequencing and quantitative time of flight mass spectrometry-mass spectrometry analyses, a 549-bp gene (lyxA) encoding D-lyxose (L-ribose) isomerase was cloned and expressed in Escherichia coli. The purified endogenous enzyme and the recombinant enzyme formed homodimers that were activated by Mn2+. C. laevoribosii D-lyxose (L-ribose) isomerase (CLLI) exhibits maximal activity at pH 6.5 and 70°C in the presence of Mn2+ for D-lyxose and L-ribose, and its isoelectric point (pI) is 4.2 (calculated pI, 4.9). The enzyme is specific for D-lyxose, L-ribose, and D-mannose, with apparent Km values of 22.4 ± 1.5 mM, 121.7 ± 10.8 mM, and 34.0 ± 1.1 mM, respectively. The catalytic efficiencies (kcat/Km) of CLLI were 84.9 ± 5.8 mM-1 s-1 for D-lyxose (V max, 5,434.8 U mg-1), 0.2 mM-1 s-1 for L-ribose (Vmax, 75.5 ± 6.0 U mg-1), and 1.4 ± 0.1 mM-1 s-1 for D-mannose (Vmax, 131.8 ± 7.4 U mg-1). The ability of lyxA to permit E. coli cells to grow on D-lyxose and L-ribose and homology searches of other sugar-related enzymes, as well as previously described sugar isomerases, suggest that CLLI is a novel type of rare sugar isomerase.

AB - A newly isolated bacterium, Cohnella laevoribosii RI-39, could grow in a defined medium with L-ribose as the sole carbon source. A 21-kDa protein isomerizing L-ribose to L-ribulose, as well as D-lyxose to D-xylulose, was purified to homogeneity from this bacterium. Based on the N-terminal and internal amino acid sequences of the purified enzyme obtained by N-terminal sequencing and quantitative time of flight mass spectrometry-mass spectrometry analyses, a 549-bp gene (lyxA) encoding D-lyxose (L-ribose) isomerase was cloned and expressed in Escherichia coli. The purified endogenous enzyme and the recombinant enzyme formed homodimers that were activated by Mn2+. C. laevoribosii D-lyxose (L-ribose) isomerase (CLLI) exhibits maximal activity at pH 6.5 and 70°C in the presence of Mn2+ for D-lyxose and L-ribose, and its isoelectric point (pI) is 4.2 (calculated pI, 4.9). The enzyme is specific for D-lyxose, L-ribose, and D-mannose, with apparent Km values of 22.4 ± 1.5 mM, 121.7 ± 10.8 mM, and 34.0 ± 1.1 mM, respectively. The catalytic efficiencies (kcat/Km) of CLLI were 84.9 ± 5.8 mM-1 s-1 for D-lyxose (V max, 5,434.8 U mg-1), 0.2 mM-1 s-1 for L-ribose (Vmax, 75.5 ± 6.0 U mg-1), and 1.4 ± 0.1 mM-1 s-1 for D-mannose (Vmax, 131.8 ± 7.4 U mg-1). The ability of lyxA to permit E. coli cells to grow on D-lyxose and L-ribose and homology searches of other sugar-related enzymes, as well as previously described sugar isomerases, suggest that CLLI is a novel type of rare sugar isomerase.

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