1
030
Y. KANEDA et al.
the pathway of M. luteolum YK-1 is under way.
aminophenazone chromogenic system in direct
enzymic assay of uric acid in serum and urine. Clin.
Chem., 26, 227–231 (1980).
Bradford, M. M., A rapid and sensitive method for
the quantitation of microgram quantities of protein
utilizing the principle of protein-dye binding. Anal.
Chem., 72, 248–254 (1976).
Laemmli, U. K., Cleavage of structural proteins
during the assembly of the head of bacteriophage T4.
Nature, 227, 680–685 (1970).
A PN 4-oxidase-encoding gene was found on a
chromosomal DNA of M. luteolum YK-1. The en-
zyme showed a high similarity to the mll6785 gene
product (a probable dehydrogenase) of a nitrogen-
ˆxing symbiotic bacterium, M. loti. The complete ge-
nome structure, including two megaplasmids, of M.
8
9
)
)
1
6)
loti has been reported. The mll6785 gene was found
at coordinates of 5585874–5584915 (a complement
direction) of a chromosomal DNA of the bacterium.
Predicted functions of other genes around the
mll6785 gene were examined. Interestingly, the mlr
1
0) Saito, H. and Miura, K., Preparation of transform-
ing deoxyribonucleic acid by phenol treatment.
Biochim. Biophys. Acta, 72, 619–629 (1963).
6
788 gene coordinated at 5587739–5588875 (a direct
11) Nakano, M., Morita, T., Yamamoto, T., Sano, H.,
Ashiuchi, M., Masui, R., Kuramitsu, S., and Yagi,
T., Puriˆcation, molecular cloning, and catalytic
activity of Schizosaccharomyces pombe pyridoxal
reductase. A possible additional family in the aldo-
keto reductase superfamily. J. Biol. Chem., 274,
direction) coded 2-methyl-3-hydroxypyridine-5-
carboxylic acid oxygenase (EC 1.14.12.4) which
catalyzes the 7th reaction step in PN degradation by
pathway I. The oxygenase has been cloned from
2
2)
Pseudomonas MA-1. The results suggested that the
probable dehydrogenase is indeed PN 4-oxidase and
M. loti has a PN degradation pathway I. More stu-
dies are needed to identify the relationship between
the nitrogen-ˆxing function and the presence of the
PN degradation pathway.
2
3185–23190 (1999).
1
2) Yagi, T., Tanouchi, A., and Hiraoka, Y., Growth
phase-dependent active transport of pyridoxine in a
ˆssion yeast, Schizosaccharomyces pombe. FEMS
Microbial. Lett., 161, 145–150 (1998).
13) Carmine, T. C., Bruchelt, G., Hahn, T., and
Nielthammer, D., Measurement of endogenous and
TNF
SK-N-SH neuroblastoma cells with an enhanced
chemiluminescence assay. J. Biolumin. Chemilumin.
, 267–272 (1994).
4) Tsuge, H. and Nakanishi, Y., Activity staining for
avoprotein oxidase. Methods Enzymol., 66, 344–350
1980).
2 2
a-mediated H O production in supernatants of
Acknowledgments
,
We acknowledge Dr. Esmond E. Snell for critical
reading of the manuscript and helpful suggestions,
and Dr. T. Taniguchi and K. Morisawa, Kochi Medi-
cal School, for protein sequence analyses.
9
1
1
1
‰
(
5) Davis, B. J., Method and application to human se-
References
rum proteins. Ann. NY. Acad. Sci., 121, 404–427
(
1964).
1
2
3
4
5
)
)
)
)
)
Snell, E. E. and Haskell, B. E., The metabolism of
6) Kaneko, T., Nakamura, Y., Sato, S., Asamizu, E.,
Kato, T., Sasamoto, S., Watanabe, A., Idesawa, K.,
Ishikawa, A., Kawashima, K., Kimura, T., Kishida,
Y., Kiyokawa, C., Kohara, M., Matsumoto, M.,
Matsuno, A., Mochizuki, Y., Nakayama, S.,
Nakazaki, N., Shimpo, S., Sugimoto, M., Takeuchi,
C., Yamada, M., and Tabata, S., Complete genome
structure of the nitrogen-ˆxing symbiotic bacterium
Mesorhizobium loti. DNA Res., 7, 381–406 (2000).
7) White, O., Eisen, J. A., Heidelberg, J. F., Hickey, E.
K., Peterson, J. D., Dodson, R. J., Haft, D. H.,
Gwinn, M. L., Nelson, W. C., Richardson, D. L.,
MoŠat, K. S., Qin, H., Jiang, L., Pamphile, W.,
Crosby, M., Shen, M., Vamathevan, J. J., Lam, P.,
McDonald, L., Utterback, T., Zalewski, C.,
Makarova, K. S., Aravind, L., Daly. M. J., Minton,
K. W., Fleisschmann, R. D., Ktchum, K. A., Nelson,
K. E., Salzberg, S., Smith, H. O., Venter, J. C., and
Fraser, C. M., Genome sequence of the radioresistant
bacterium Deinococcus radiodurans R1. Science, 286,
6
vitamin B . In ``Comprehensive Biochemistry'' eds.
Florkin, M. and Stotz, E. H., Elsevier North
W
Holland, New York, vol. 21, pp. 47–67 (1971).
Nelson, M. J. and Snell, E. E., Enzymes of vitamin
B degradation. Puriˆcation and properties of 5-pyri-
6
doxic acid oxygenase from Arthrobacter sp. J. Biol.
Chem., 261, 15115–15120 (1986).
Sundaram, T. K. and Snell, E. E., The bacterial oxi-
6
dation of vitamin B . V. The enzymatic formation of
pyridoxal and isopyridoxal from pyridoxine. J. Biol.
Chem., 244, 2577–2584 (1969).
Trongpanich, Y., Abe, K., Kaneda, Y., Morita, T.,
and Yagi, T., Puriˆcation and characterization of
pyridoxal 4-dehydrogenase from Microbacterium
luteolum. Biosci. Biotechnol. Biochem., in press.
Yagi, T., Kirino, J., Yamamoto, S., and Nozaki, M.,
1
A
simple preparation method for apoaspartate
aminotransferase from Escherichia coli B, and its ap-
plication for the assay of pyridoxal and pyridoxamine
5?-phosphate. J. Biochem. (Tokyo) 98, 921–926
1
571–1577 (1999).
(
1985).
1
8) Boyd, L. A., Adam, L., Pelcher, L. E., McHughen,
A., Hirji, R., and Selvaraj, G., Characterization of
an Escherichia coli gene encoding betaine aldehyde
dehydrogenase (BADH): structural similarity to
mammalian ALDHs and a plant BADH. Gene, 103,
45–52 (1991).
6
7
)
)
Wada, H. and Snell, E. E., The enzymatic oxidation
of pyridoxine and pyridoxamine phosphate. J. Biol.
Chem., 236, 2089–2095 (1961).
Fossati, P., Principe, L., and Berti, G., Use
of 3,5-dichloro-2-hydroxybenzenesulfonic acid 4-
W