The missing link in the fungal L-arabinose catabolic pathway, identification of the L-xylulose reductase gene.
The fungal L-arabinose pathway consists of five enzymes, aldose reductase, L-arabinitol 4-dehydrogenase, L-xylulose reductase, xylitol dehydrogenase, and xylulokinase. All the genes encoding the enzymes of this pathway are known except for that of L-xylulose reductase (EC 1.1.1.10). We identified a gene encoding this enzyme from the filamentous fungus Trichoderma reesei (Hypocrea jecorina). The gene was named lxr1. It was overexpressed in the yeast Saccharomyces cerevisiae, and the enzyme activity was confirmed in a yeast cell extract. Overexpression of all enzymes of the L-arabinose pathway in S. cerevisiae led to growth of S. cerevisiae on L-arabinose; i.e., we could show that the pathway is active in a heterologous host. The lxr1 gene encoded a protein with 266 amino acids and a calculated molecular mass of 28 428 Da. The LXRI protein is an NADPH-specific reductase. It has activity with L-xylulose, D-xylulose, D-fructose, and L-sorbose. The highest affinity is toward L-xylulose (K(m) = 16 mM). In the reverse direction, we found activity with xylitol, D-arabinitol, D-mannitol, and D-sorbitol. It requires a bivalent cation for activity. It belongs to the protein family of short chain dehydrogenases. The enzyme is catalytically similar and homologous in sequence to a D-mannitol:NADP 2-dehydrogenase (EC 1.1.1.138).
Richard
Peter
P
VTT Biotechnology, P.O. Box 1500, FIN-02044 VTT, Finland. Peter.Richard@vtt.fi
Putkonen
Mikko
M
Väänänen
Ritva
R
Londesborough
John
J
Penttilä
Merja
M
D000595:Amino Acid Sequence; D001089:Arabinose / Q000378:metabolism*; D002384:Catalysis; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D005822:Genetic Vectors / Q000138:chemical synthesis; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D013401:Sugar Alcohol Dehydrogenases / Q000235:genetics* / Q000302:isolation & purification* / Q000502:physiology; D014242:Trichoderma / Q000201:enzymology* / Q000235:genetics*
D005656:Fungal Proteins; D001089:Arabinose; D013401:Sugar Alcohol Dehydrogenases; C023300:L-xylulose reductase