Kinetic analysis of mouse retinal dehydrogenase type-2 (RALDH2) for retinal substrates.
Retinal dehydrogenase (RALDH) isozymes catalyze the terminal oxidation of retinol into retinoic acid (RA) that is essential for embryogenesis and tissue differentiation. To understand the role of mouse type 2 RALDH in synthesizing the ligands (all-trans and 9-cis RA) needed to bind and activate nuclear RA receptors, we determined the detailed kinetic properties of RALDH2 for various retinal substrates. Purified recombinant RALDH2 showed a pH optimum of 9.0 for all-trans retinal oxidation. The activity of the enzyme was lower at 37 degrees C compared to 25 degrees C. The efficiency of conversion of all-trans retinal to RA was 2- and 5-fold higher than 13-cis and 9-cis retinal, respectively. The K(m) for all-trans and 13-cis retinal were similar (0.66 and 0.62 microM, respectively). However, the K(m) of RALDH2 for 9-cis retinal substrate (2.25 microM) was 3-fold higher compared to all-trans and 13-cis retinal substrates. Among several reagents tested for their ability to either inhibit or activate RALDH2, citral and para-hydroxymercuribenzoic acid (p-HMB) inhibited and MgCl(2) activated the reaction. Comparison of the kinetic properties of RALDH2 for retinal substrates and its activity towards various reagents with those of previously reported rat kidney RALDH1 and human liver aldehyde dehydrogenase-1 showed distinct differences. Since RALDH2 has low K(m) and high catalytic efficiency for all-trans retinal, it may likely be involved in the production of all-trans RA in vivo.
Gagnon
Isabelle
I
Laboratory of Nutrition and Cancer, Centre Hospitalier de l'Université de Montreal-Research Centre, Hotel-Dieu Campus, Université de Montreal, 3840 rue Saint-Urbain, Montreal, Quebec, Canada H2W 1T8.
Duester
Gregg
G
Bhat
Pangala V
PV
D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000077556:Alitretinoin; D000818:Animals; D002384:Catalysis; D003001:Cloning, Molecular; D004224:Diterpenes; D015971:Gene Expression Regulation, Enzymologic; D006863:Hydrogen-Ion Concentration; D015474:Isotretinoin / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D011994:Recombinant Proteins / Q000737:chemistry; D050697:Retinal Dehydrogenase; D012172:Retinaldehyde / Q000378:metabolism; D013696:Temperature; D014212:Tretinoin / Q000378:metabolism; D014801:Vitamin A / Q000378:metabolism
D004224:Diterpenes; D011994:Recombinant Proteins; D014801:Vitamin A; D000077556:Alitretinoin; C031338:13-cis-retinal; C031390:9-cis-retinal; D014212:Tretinoin; D000445:Aldehyde Oxidoreductases; D050697:Retinal Dehydrogenase; D015474:Isotretinoin; D012172:Retinaldehyde