Identification of a new tetrameric pyridoxal 4-dehydrogenase as the second enzyme in the degradation pathway for pyridoxine in a nitrogen-fixing symbiotic bacterium, Mesorhizobium loti.
We have found for the first time that a chromosomal gene, mlr6807, in Mesorhizobium loti encodes a new tetrameric pyridoxal 4-dehydrogenase (PLDH). The recombinant enzyme expressed in Escherichia coli cells was homogenously purified and characterized. The enzyme consisted of four subunits each with a molecular weight of 26,000+/-1000, and exhibited Km and kcat values of 91+/-2 microM and 149+/-1s(-1), respectively. PLDH used NAD+ as a cosubstrate, showed no activity toward sugars, and belonged to a short-chain dehydrogenase/reductase family. The mlr6807 gene-disrupted M. loti cells could grow in a nutrient-rich TY medium but not in a synthetic one containing pyridoxine or pyridoxamine as the sole carbon and nitrogen source. Thus, it was found that PLDH is essential for the assimilation of vitamin B6 compounds and the second step enzyme in their degradation pathway in M. loti.
Yokochi
Nana
N
Department of Bioresources Science, Faculty of Agriculture, Kochi University, Monobe-Otsu 200, Nankoku, Kochi 783-8502, Japan.
Nishimura
Saki
S
Yoshikane
Yu
Y
Ohnishi
Kouhei
K
Yagi
Toshiharu
T
D000429:Alcohol Oxidoreductases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D020561:Alphaproteobacteria / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002244:Carbon / Q000378:metabolism; D004789:Enzyme Activation; D004926:Escherichia coli / Q000166:cytology / Q000201:enzymology / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D009584:Nitrogen / Q000378:metabolism*; D011733:Pyridoxamine / Q000378:metabolism; D011736:Pyridoxine / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013559:Symbiosis; D013816:Thermodynamics; D025101:Vitamin B 6 / Q000378:metabolism*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D011733:Pyridoxamine; D002244:Carbon; D025101:Vitamin B 6; D000429:Alcohol Oxidoreductases; C042642:pyridoxal dehydrogenase; D011736:Pyridoxine; D009584:Nitrogen