Identification in the mold Hypocrea jecorina of the first fungal D-galacturonic acid reductase.
A d-galacturonic acid reductase and the corresponding gene were identified from the mold Hypocrea jecorina (Trichoderma reesei). We hypothesize that the enzyme is part of a fungal d-galacturonic acid catabolic pathway which has not been described previously and which is distinctly different from the bacterial pathway. H. jecorina grown on d-galacturonic acid exhibits an NADPH-dependent d-galacturonic acid reductase activity. This activity is absent when the mold is grown on other carbon sources. The d-galacturonic acid reductase was purified, and tryptic digests of the purified protein were sequenced. The open reading frame of the corresponding gene was then cloned from a cDNA library. The open reading frame was functionally expressed in the yeast Saccharomyces cerevisiae. A histidine-tagged protein was purified, and the enzyme kinetics were characterized. The enzyme converts in a reversible reaction from d-galacturonic acid and NADPH to l-galactonic acid and NADP. The enzyme also exhibits activity with d-glucuronic acid and dl-glyceraldehyde.
Kuorelahti
Satu
S
VTT Biotechnology, P.O. Box 1500, 02044 VTT, Espoo, Finland.
Kalkkinen
Nisse
N
Penttilä
Merja
M
Londesborough
John
J
Richard
Peter
P
D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D015723:Gene Library; D006603:Hexuronic Acids / Q000737:chemistry* / Q000378:metabolism; D035901:Hypocrea / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D064430:NAD (+) and NADP (+) Dependent Alcohol Oxidoreductases; D009249:NADP / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013379:Substrate Specificity / Q000502:physiology; D013400:Sugar Acids / Q000737:chemistry / Q000378:metabolism
D005656:Fungal Proteins; D006603:Hexuronic Acids; D013400:Sugar Acids; C012991:galactonic acid; C007819:galacturonic acid; D009249:NADP; D000429:Alcohol Oxidoreductases; D064430:NAD (+) and NADP (+) Dependent Alcohol Oxidoreductases; C477073:D-galacturonate reductase