Identification of a mouse short-chain dehydrogenase/reductase gene, retinol dehydrogenase-similar. Function of non-catalytic amino acid residues in enzyme activity

J Biol Chem. 2003 Oct 10;278(41):40079-87. doi: 10.1074/jbc.M304910200. Epub 2003 Jul 10.


We report a mouse short-chain dehydrogenase/reductase (SDR), retinol dehydrogenase-similar (RDH-S), with intense mRNA expression in liver and kidney. The RDH-S gene localizes to chromosome 10D3 with the SDR subfamily that catalyzes metabolism of retinoids and 3 alpha-hydroxysteroids. RDH-S has no activity with prototypical retinoid/steroid substrates, despite 92% amino acid similarity to mouse RDH1. This afforded the opportunity to analyze for functions of non-catalytic SDR residues. We produced RDH-S Delta 3 by mutating RDH-S to remove an "additional" Asn residue relative to RDH1 in its center, to convert three residues into RDH1 residues (L121P, S122N, and Q123E), and to substitute RDH1 sequence G208FKTCVTSSD for RDH-S sequence F208-FLTGMASSA. RDH-S Delta 3 catalyzed all-trans-retinol and 5 alpha-androstane-3 alpha,17 alpha-diol (3 alpha-adiol) metabolism 60-70% as efficiently (Vm/Km) as RDH1. Conversely, substituting RDH-S sequence F208FLTGMASSA into RDH1 produced a chimera (viz. C3) that was inactive with all-trans-retinol, but was 4-fold more efficient with 3 alpha-adiol. A single RDH1 mutation in the C3 region (K210L) reduced efficiency for all-trans-retinol by >1250-fold. In contrast, the C3 area mutation C212G enhanced efficiency with all-trans-retinol by approximately 2.4-fold. This represents a >6000-fold difference in catalytic efficiency for two enzymes that differ by a single non-catalytic amino acid residue. Another chimera (viz. C5) retained efficiency with all-trans-retinol, but was not saturated and was weakly active with 3 alpha-adiol, stemming from three residue differences (K224Q, K229Q, and A230T). The residues studied contribute to the substrate-binding pocket: molecular modeling indicated that they would affect orientation of substrates with the catalytic residues. These data report a new member of the SDR gene family, provide insight into the function of non-catalytic SDR residues, and illustrate that limited changes in the multifunctional SDR yield major alterations in substrate specificity and/or catalytic efficiency.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alcohol Oxidoreductases / chemistry
  • Alcohol Oxidoreductases / genetics*
  • Alcohol Oxidoreductases / metabolism*
  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Base Sequence
  • Binding Sites
  • Chromosome Mapping
  • Cloning, Molecular
  • DNA, Complementary / genetics
  • Fatty Acid Synthases / chemistry
  • Fatty Acid Synthases / genetics*
  • Fatty Acid Synthases / metabolism*
  • Kidney / enzymology
  • Kinetics
  • Liver / enzymology
  • Mice
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • NADH, NADPH Oxidoreductases / chemistry
  • NADH, NADPH Oxidoreductases / genetics*
  • NADH, NADPH Oxidoreductases / metabolism*
  • Protein Conformation
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Substrate Specificity


  • DNA, Complementary
  • RNA, Messenger
  • Recombinant Fusion Proteins
  • Alcohol Oxidoreductases
  • retinol dehydrogenase
  • short chain trans-2-enoyl-CoA reductase
  • NADH, NADPH Oxidoreductases
  • Fatty Acid Synthases

Associated data

  • GENBANK/AY046408