Novel reactions of L-rhamnose isomerase from Pseudomonas stutzeri and its relation with D-xylose isomerase via substrate specificity.
Escherichia coli strain JM 109 harboring 6 x His-tag L-rhamnose isomerase (L-RhI) from Pseudomonas stutzeri allowed a 20-fold increase in the volumetric yield of soluble enzyme compared to the value for the intrinsic yield. Detailed studies on the substrate specificity of the purified His-tagged protein revealed that it catalyzed previously unknown common and rare aldo/ketotetrose, aldo/ketopentose, and aldo/ketohexose substrates in both D- and L-forms, for instance, erythrose, threose, xylose, lyxose, ribose, glucose, mannose, galactose, altrose, tagatose, sorbose, psicose, and fructose. Using a high enzyme-substrate ratio in extended reactions, the enzyme-catalyzed interconversion reactions from which two different products from one substrate were formed: L-lyxose, L-glucose, L-tagatose and D-allose were isomerized to L-xylulose and L-xylose, L-fructose and L-mannose, L-galactose and L-talose, and D-psicose and D-altrose, in that order. Kinetic studies, however, showed that L-rhamnose with Km and Vmax values of 11 mM and 240 U/mg, respectively, was the most preferred substrate, followed by L-mannose, L-lyxose, D-ribose, and D-allose. Based on the observed catalytic mode of action, these new findings reflected a hitherto undetected interrelation between L-RhI and D-xylose isomerase (D-XI).
Leang
Khim
K
Department of Biochemistry and Food Science, Faculty of Agriculture and Rare Sugar Research Center, Kagawa University, Ikenobe 2393, Miki-cho, Kagawa 761-0795, Japan.
Takada
Goro
G
Fukai
Yoshinori
Y
Morimoto
Kenji
K
Granström
Tom Birger
TB
Izumori
Ken
K
D019747:Aldose-Ketose Isomerases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D044223:Pseudomonas stutzeri / Q000201:enzymology*; D016415:Sequence Alignment; D013302:Streptomyces / Q000201:enzymology; D013379:Substrate Specificity
D001426:Bacterial Proteins; D002241:Carbohydrates; D008670:Metals; D019747:Aldose-Ketose Isomerases; C062878:L-rhamnose isomerase; C023298:xylose isomerase