Isocitrate Dehydrogenases and NADP‐Alcohol Dehydrogenase of Euglena gracilis var. bacillaris*

Abstract

SYNOPSIS. Nicotinamide adenine dinucleotide phosphate (NADP) and nicotinamide adenine dinucleotide (NAD) linked isocitrate dehydrogenase and NADP linked alcohol dehydrogenase have been detected in Euglena gracilis var. bacillaris. The NADP isocitrate dehydrogenase showed half‐maximal activity at a concentration of 3 × 10−5 M DL‐isocitrate, but did not follow simple Michaelis‐Menten kinetics with respect to substrate concentration. The optimal NADP concentration was about 0.06 mM, and activity fell off sharply on either side of this optimum. Fresh preparations of the enzyme migrated as single bands in disc electrophoresis, but two enzymatically active bands were present after frozen storage. The NAD isocitrate dehydrogenase followed Michaelis‐Menten kinetics with respect to substrate. In crude extracts, no requirement for adenosine monophosphate, adenosine diphosphate, or sulfhydryl compounds could be found. NADP alcohol dehydrogenase activity could be found with either ethanol or propanol as substrate. Low concentrations of coenzyme A were moderately inhibitory. In tris(hydroxymethyl) aminomethane buffer (tris buffer), Euglena extracts reduced NAD slowly in the absence of exogenous substrate. In the absence of tris, no such reduction occurred. A similar phenomenon was observed with NADP.

Gene Cloning and Biochemical Characterization of an Alcohol Dehydrogenase from Euglena gracilis1

“…Early reports have documented NAD + (Hurlbert and Rittenberg 1962; Mego and Farb 1974) and NADP + –ADH activities (Danforth and Hunter 1966; Eshleman and Danforth 1964; Wilson and Danforth 1958) in E. gracilis grown on ethanol as carbon source. In addition, a microsomal ethanol‐oxidizing system (cytochrome P‐450) was also found in E. gracilis grown on lactate (Julistiono and Briand 1992).…”

Subcellular Location and Some Properties of Isocitrate Dehydrogenase Isozymes inEuglena gracilis

“…Z3 ) Euglena NAD+ -linked enzyme, however, was insensitive to adenine nucleotides, similarly to that of pea mitochondria. z4 ) Danforth and Hunter 1 ) and Flavell and Dart 14 ) reported with no data that the activity ofNAD+ -linked enzyme was inhibited by ATP, ADP and AMP.…”

“…1.41] and the other NADP+ [threo-os-isocitrate: NADP+ oxidoreductase (decarboxylating), EC 1.1.1.42].1) Previously no suitable technique has been devised for obtaining an adequate cell homogenate of E. gracilis, and accordingly subcellular location and detailed properties of these isozymes in this protozoan have not been studied extensively. Since the isozymes probably have important functions controlling the carbon flow between the tricarboxylic acid cycle and the glyoxylate cycle and supplying 2-oxoglutarate in Euglena, their locations and properties are studied in the present paper.…”