title: Molecular characterization of an aldo-keto reductase from Marivirga tractuosa that converts retinal to retinol.
abstract: A recombinant aldo-keto reductase (AKR) from Marivirga tractuosa was purified with a specific activity of 0.32unitml(-1) for all-trans-retinal with a 72kDa dimer. The enzyme had substrate specificity for aldehydes but not for alcohols, carbonyls, or monosaccharides. The enzyme turnover was the highest for benzaldehyde (kcat=446min(-1)), whereas the affinity and catalytic efficiency were the highest for all-trans-retinal (Km=48μM, kcat/Km=427mM(-1)min(-1)) among the tested substrates. The optimal reaction conditions for the production of all-trans-retinol from all-trans-retinal by M. tractuosa AKR were pH 7.5, 30°C, 5% (v/v) methanol, 1% (w/v) hydroquinone, 10mM NADPH, 1710mgl(-1) all-trans-retinal, and 3unitml(-1) enzyme. Under these optimized conditions, the enzyme produced 1090mgml(-1) all-trans-retinol, with a conversion yield of 64% (w/w) and a volumetric productivity of 818mgl(-1)h(-1). AKR from M. tractuosa showed no activity for all-trans-retinol using NADP(+) as a cofactor, whereas human AKR exhibited activity. When the cofactor-binding residues (Ala158, Lys212, and Gln270) of M. tractuosa AKR were changed to the corresponding residues of human AKR (Ser160, Pro212, and Glu272), the A158S and Q270E variants exhibited activity for all-trans-retinol. Thus, amino acids at positions 158 and 270 of M. tractuosa AKR are determinant residues of the activity for all-trans-retinol.
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Republic of Korea.
authors.forename: Seung-Hye
authors.initials: SH
authors.lastname: Hong
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Republic of Korea.
authors.forename: Hyun-Koo
authors.initials: HK
authors.lastname: Nam
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Republic of Korea.
authors.forename: Kyoung-Rok
authors.initials: KR
authors.lastname: Kim
authors.affiliation: Division of Applied Life Science, EB-NCRC and PMBBRC, Gyeongsang National University, Jinju 660-701, Republic of Korea.
authors.forename: Seon-Won
authors.initials: SW
authors.lastname: Kim
authors.affiliation: Department of Bioscience and Biotechnology, Konkuk University, Seoul 143-701, Republic of Korea. Electronic address: deokkun@konkuk.ac.kr.
authors.forename: Deok-Kun
authors.initials: DK
authors.lastname: Oh
mesh_terms: D000449:Aldehyde Reductase / Q000235:genetics / Q000378:metabolism*; D000074408:Aldo-Keto Reductases; D041963:Bacteroidetes / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D012172:Retinaldehyde / Q000737:chemistry / Q000378:metabolism*; D013379:Substrate Specificity; D014801:Vitamin A / Q000737:chemistry / Q000378:metabolism*
chemical_list: D011994:Recombinant Proteins; D014801:Vitamin A; D000074408:Aldo-Keto Reductases; D000449:Aldehyde Reductase; D012172:Retinaldehyde
keywords: Aldo-keto reductase; Cofactor-binding residues; Marivirga tractuosa; Retinal; Retinol