Correlation Between L-Rhamnose Isomerase and D-Xylose Isomerase

OUTLINE OF D-ALLOSEMASS-PRODUCTION

1Kenji Morimoto*, 2Motofumi Ozaki, 2Chang-Su Park, 3Kei Takeshita, 2Tsuyoshi Shimonishi, 4Kyosuke Yamada, 1Tom Birger Granström, 1Goro Takata, 1Masaaki TokudaAND1Ken Izumori

1RareSugarResearchCenter, Kagawa University, Miki-cho, Kagawa 761-0795, Japan.

2Kagawa IndustrySupportFoundation, Hayashi-cho, Takamatsu, Kagawa 761-0301,Japan.3Fushimi Pharmaceutical Co., Ltd, Nakatsu-cho Marugame, Kagawa, 763-8605, Japan.4ORGANO Corporation, Shinsuna Koutou-ku, Tokyo,136-8631, Japan.

Raresugars are monosaccharides, whichexistrare in nature. It is knownthat the somehexoseraresugars havesomephysiologicallyactivefunctions. One of them, D-allose, has abilities to suppresscancertissuegrowth and protect ischemia. We reported aboutD-psicose massproduction in InternationalSymposiumRareSugar in 2002. We establishedmassproduction of D-allosefromD-psicose by usingmicrobial enzymaticreaction.

L-Rhamnoseisomerase, L-rhamnose ketol-isomerase [EC 5.3.1.14], whichisomerizes L-rhamnose to L-rhamnulose, has beenfound in Escherichia coli, Salmonella, and Pseudomonas. L-RhamnoseisomerasefromPseudomonas stutzeri was reported to catalyzeisomerizations betweenD-allose - D-psicose besidesL-rhamnose - L-rhamnulose. Moreover, the enzyme was then sucsessfullycloned into E. coli and nucleotidesequence was elucidated.

The recombinantE. coli was cultivated in jar-fermenter. Aftercelldisruption by an ultrasonicator, cellfreeextract was immobilized to ChitopearlBCW-2510. A bioreactor was constructed and 50% (w/v) D-psicose was continuouslyconvertedintoD-allose at 42˚C. The reactionequilibrium was D-psicose (70): D-allose (30) in thiscase. Park et. al. publishresults in detail at thissymposium. The reactionmixture was continuouslyseparatedD-allose and D-psicose by simulatedmoving bed system using TREZONE (ORGANO Corp). Ozaki et. al. publishdetailedconditions and results at thissymposium. D-allosefraction was concentrated by evaporatoruntil ca. 50% (w/v) and keeping at roomtemperatureallowed syrup to grow D-allosecrystalwithoutseedcrystal. The yield of D-allosecrystal was 50% of D-psicose which was used in reaction.