Aldehyde dehydrogenase (CoA-acetylating) and the mechanism of ethanol formation in the amitochondriate protist, Giardia lamblia.
The so far unelucidated pathway of formation of ethanol, one of the major end products of the fermentative metabolism of the amitochondriate protist, Giardia lamblia, was examined. Two NAD-dependent enzymatic activities, an acetaldehyde dehydrogenase (CoA-acetylating) (EC 1.2.1.10) and an alcohol dehydrogenase (EC 1.1.1.1) were detected. These are assumed to catalyze the formation of ethanol from acetyl-CoA via acetaldehyde. The first activity, present on a 95-kDa protein, was purified. It catalyzed the reversible interconversion of acetyl-CoA to acetaldehyde and CoA-SH with NAD but not NADP as cofactor. In the direction of aldehyde formation acetyl-CoA was the preferred substrate. Propionyl-CoA and isobutyryl-CoA were reduced with lower efficiency while succinyl-CoA and benzoyl-CoA were not. In the direction of acyl-CoA formation, acetaldehyde was the preferred substrate. Propionaldehyde and isobutyraldehyde were utilized at a lower efficiency while formaldehyde, benzaldehyde, and acetone were not. The second activity, a primary alcohol dehydrogenase, was also NAD-specific and used preferentially ethanol as substrate. Sequencing data of peptides from the purified protein and Northern and Southern analysis indicated that the same polypeptide, which belongs to the bifunctional aldehyde/alcohol dehydrogenase enzyme family, carried both activities. These activities define the pathway to ethanol in G. lamblia as a two step-processes: (i) acetyl-CoA + NADH<-->acetaldehyde + CoA-SH + NAD+ and (ii) acetaldehyde + NADH<-->ethanol + NAD+. In contrast to most eukaryotes in which ethanol formation proceeds from pyruvate via acetaldehyde, the G. lamblia pathway departs from acetyl-CoA, a more distal product of extended glycolysis.
Sánchez
L B
LB
Rockefeller University, New York, New York 10021, USA. sanchel@rockvax.rockefeller.edu
D000079:Acetaldehyde / Q000378:metabolism; D000426:Alcohol Dehydrogenase / Q000378:metabolism; D000445:Aldehyde Oxidoreductases / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D004789:Enzyme Activation; D000431:Ethanol / Q000378:metabolism*; D016829:Giardia lamblia / Q000201:enzymology* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D013347:Subcellular Fractions / Q000378:metabolism
D000431:Ethanol; D000426:Alcohol Dehydrogenase; D000445:Aldehyde Oxidoreductases; C027014:acetaldehyde dehydrogenase (acylating); D000079:Acetaldehyde