Cofactor requirements of the L-malate dehydrogenase of Pseudomonas ovalis Chester.
1. The l-malate dehydrogenase of Pseudomonas ovalis Chester, which is independent of nicotinamide nucleotides and which is structurally and functionally bound to the cell-wall membrane, has been prepared in a soluble form and partially purified. 2. The purified dehydrogenase exhibits a triple cofactor requirement for FAD, quinone and phospholipid, and in the presence of these cofactors can utilize 2,6-dichlorophenol-indophenol as hydrogen acceptor. 3. The formation of reduced forms of FAD was not detected, but in the presence of both FAD and phospholipid the enzyme catalysed the reduction of quinone by l-malate at rates equivalent to those obtained with 2,6-dichlorophenol-indophenol as terminal acceptor. The l-malate dehydrogenase of Ps. ovalis Chester is therefore an l-malate-quinone oxidoreductase. 4. The quinone and the phospholipids present in the fragments of the cell-wall membrane from which the soluble dehydrogenase was prepared have been extracted and purified. The quinone was identified as coenzyme Q(9). At least eight phospholipids were detected, and the major component is an unsaturated phosphatidylethanolamine. 5. The nature of the phospholipid required to activate the enzyme depends on the nature of the quinone used in the assay system. When 2-methyl-1,4-naphthaquinone is used, a wide variety of phospholipids, including all those isolated from the organism, will activate the enzyme, but when coenzyme Q(9) is used the phospholipid specificity of the enzyme is much more restricted, and the most effective activator is the unsaturated phosphatidylethanolamine isolated from the organism. 6. Evidence is presented to support the view that the restricted phospholipid specificity exhibited by the enzyme in the presence of coenzyme Q(9), as opposed to the broad specificity exhibited when 2-methyl-1,4-naphthaquinone is used, is due to the fact that coenzyme Q(9) has a large substituent on position 3.
D002845:Chromatography; D002855:Chromatography, Thin Layer; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism*; D008291:Malate Dehydrogenase / Q000032:analysis* / Q000378:metabolism*; D009285:Naphthoquinones / Q000378:metabolism*; D010714:Phosphatidylethanolamines / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology*; D014451:Ubiquinone / Q000378:metabolism; D014812:Vitamin K / Q000378:metabolism
D009285:Naphthoquinones; D010714:Phosphatidylethanolamines; D014812:Vitamin K; D014451:Ubiquinone; D005182:Flavin-Adenine Dinucleotide; D008291:Malate Dehydrogenase