Pyrroloquinoline quinone-dependent dehydrogenases from Ketogulonicigenium vulgare catalyze the direct conversion of L-sorbosone to L-ascorbic acid.
A novel enzyme, L-sorbosone dehydrogenase 1 (SNDH1), which directly converts L-sorbosone to L-ascorbic acid (L-AA), was isolated from Ketogulonicigenium vulgare DSM 4025 and characterized. This enzyme was a homooligomer of 75-kDa subunits containing pyrroloquinoline quinone (PQQ) and heme c as the prosthetic groups. Two isozymes of SNDH, SNDH2 consisting of 75-kDa and 55-kDa subunits and SNDH3 consisting of 55-kDa subunits, were also purified from the bacterium. All of the SNDHs produced L-AA, as well as 2-keto-L-gulonic acid (2KGA), from L-sorbosone, suggesting that tautomerization of L-sorbosone causes the dual conversion by SNDHs. The sndH gene coding for SNDH1 was isolated and analyzed. The N-terminal four-fifths of the SNDH amino acid sequence exhibited 40% identity to the sequence of a soluble quinoprotein glucose dehydrogenase from Acinetobacter calcoaceticus. The C-terminal one-fifth of the sequence exhibited similarity to a c-type cytochrome with a heme-binding motif. A lysate of Escherichia coli cells expressing sndH exhibited SNDH activity in the presence of PQQ and CaCl2. Gene disruption analysis of K. vulgare indicated that all of the SNDH proteins are encoded by the sndH gene. The 55-kDa subunit was derived from the 75-kDa subunit, as indicated by cleavage of the C-terminal domain in the bacterial cells.
Miyazaki
Taro
T
Department of Applied Microbiology, Nippon Roche Research Center (NRRC), Kamakura, Kanagawa 247-8530, Japan. miyazakitru@chugai-pharm.co.jp
Sugisawa
Teruhide
T
Hoshino
Tatsuo
T
D000445:Aldehyde Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001205:Ascorbic Acid / Q000378:metabolism*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D006418:Heme / Q000737:chemistry; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016254:Mutagenesis, Insertional; D045542:PQQ Cofactor / Q000737:chemistry; D021122:Protein Subunits; D041902:Rhodobacteraceae / Q000201:enzymology* / Q000235:genetics; D013013:Sorbose / Q000031:analogs & derivatives* / Q000378:metabolism
D004269:DNA, Bacterial; D007527:Isoenzymes; D021122:Protein Subunits; C013686:sorbosone; D006418:Heme; D045542:PQQ Cofactor; D000445:Aldehyde Oxidoreductases; C095707:L-sorbosone dehydrogenase; D013013:Sorbose; D001205:Ascorbic Acid