COENZYME SPECIFICITY OF D-AMINO ACID OXIDASE FOR THE ADENYLATE MOIETY OF FAD.

Abstract

Analogues of FAD have been synthesized from pyridinium FMN and 4-morpholine N , N ′-dicyclohexylcarboxamidinium nucleoside 5′-phosphoromorpholidates in pyridane. The similar spectral and chromatographic properties of the analogues are described. The specific reactivity of the adenylate portion of FAD has been delineated by the following observations on the behavior of adenosyl-replaced analogues of FAD and of AMP with D -amino acid oxidase ( D -amino acid:O 2 oxidoreductase (deaminating), EC 1.4.3.3): 1. 1. Substitution of deoxyribofuranosyl for the ribofuranosyl portion of FAD results in a large decrease in coenzymatic activity. FdeAD has a K m of 4.5·10 −6 M and in 10-fold larger amounts is only 52% as active as FAD with a K m of 4.5·10 −7 M. 2. 2. All other replacements of the adenosyl moiety of FAD by common purine and pyrimidine ribo- and deoxyribofuranosyl portions yield analogues which are active only very weakly either as coenzymes or competitive inhibitors of FAD. 3. 3. A further reflection of the propensity for binding of the adenylate portion of FAD is seen in the decreased ability of nucleoside 5′-monophosphates other than AMP to inhibit attachment of FAD as coenzyme. Overall, the results indicate that D -amino acid oxidase exhibits a consederable degree of coenzyme specificity with regard to the 6-aminopurine ribofuranosyl or AMP portion of FAD.