Studies with Analogues of Nicotinamide Adenine Dinucleotide

Abstract

Through a series of correlative studies, the stereochemistry of the enzymatic processes with the coenzyme analogues of NAD+, 3-acetylpyridine-adenine dinucleotide, thionicotinamide-adenine dinucleotide and 3-cyanopyridine-adenine dinucleotide, have been examined for horse liver and yeast alcohol dehydrogenase (both A-specific for NAD+ ) and glutamate dehydrogenase (Bspecific for NAD+). I n each case the stereochemistry of the process with respect to substrate and coenzyme remains identical to that for NAD+. The implications for this are examined in view of the different geometry of the substituents a t the 3-position of the pyridine ring. A ready conversion of the thionicotinamide analogue of NADH to NADH is also described.Through the early pioneering work of Westheimer et al., the stereospecificity of the NAD+ or NADP+-dependent dehydrogenases for both the coenzyme and the substrate was demonstrated [1 to 31. I n particular it was shown that such enzymes could be classified into two distinct groups A and B depending on which side of the pyridinium ring, hydrogen transfer occurs [4,5]. I n a most elegant series of experiments by Cornforth et al., it was clearly demonstrated that the A-group enzymes abstracted the pro-R hydrogen a t C-4 of the dihydropyridine ring of NADH and the B-group enzymes the pro-S hydrogen [6,7].The stereochemistry of the hydrogen abstracted from C-1 of ethanol by the liver and the yeast alcohol dehydrogenases has been demonstrated as pro-R [8].A number of active analogues of NAD+ (NADP+) where the nicotinamide is replaced by a pyridine substituted a t C-3 with an electron-attracting group, for example thionicotinamide adenine dinucleotide (sNAD+) [9-111, selenonicotinamide adenine dinucleotide phosphate [ 121 and 3-acetylpyridine adenine dinucleotide (acNAD+) [13,14], have been prepared.However, as yet no study has been undertaken to determine whether such analogues posses the same Abbreviations. sNAD, thionicotinamide adenine dinucleotide; acNAD, 3-acetylpyridine adenine dinucleotide; cnNAD, 3-cyanopyridine adenine dinucleotide ; ORD, optical rotatory dispersion.Enzymes. NAD(P) nncleosidase (EC 3.2.2.6) ; alcohol dehydrogenase (EC 1.1.1.1) ; L-glutamate dehydrogenase (EC 1.4.1.2). stereochemical course as that for NAD+. Clearly is has been assumed that the similar geometry of the substituents a t C-3 of the pyridinium ring would result in an identical stereochemical reaction course. Recently we described the preparation of a new active analogue of NAD+, 3-cyanopyridine adenine dinucleotide (cnNAD+) [15]. Since this is the first analogue to be prepared with a linear functional group a t C-3 and since we have a t our disposal a ready means of correlation between sNADH and cnNADH by simple reaction with silver nitrate, we considered it informative to study the relative stereochemistry of the hydrogen transfer for these analogues. METHODS AND RESULTS MATERIALS AND GENERAL INDICATIONSNAD+ labeled grade 111, approx. 98O/, pure, pig's brain NAD( P) nucleosidase and the two dehydrogenases, hor...

Nicotinamide 3,N4-ethenocytosine dinucleotide, an analog of nicotinamide adenine dinucleotide. Synthesis and enzyme studies

“…L he prominent biological roles played by nicotinamide adenine dinucleotide (NAD+) (1) and its reduced form NADH have generated considerable interest in these compounds and in the enzymes which utilize them as coenzymes or substrates. One investigative technique that has been employed in determining structure and function relationships of these compounds has been the preparation of NAD+ analogs and the study of their interaction with dehydrogenases and other enzymes (Sund, 1968a; Colowick, et al, 1966;Biellmann et al, 1974;Chaykin, 1967;Suhadolnik et al, 1974). Analogs have also been used in studies of the mechanisms of dehydrogenase action (Sund, 1968b), the detection of heterogeneity of enzymes having the same function (Kaplan, 1963a), and the evolution of enzyme structure From the Department of Biochemistry, School of Chemical Sciences, and School of Basic Medical Sciences, University of Illinois, Urbana, Illinois 61801.…”

Mechanisms of zinc ion catalysis in small molecules and enzymes

“…In contrast, coenzyme specificity is quite restrictive. The early studies of Anderson et al (53, 5d) and more recently of Biellmann et al (55) indicate that the nicotinamide ring can be chemically altered only at the 3-position without destroying the chemical functionality of the dinucleotide. Furthermore, the activity of coenzyme analogs chemically altered at the 3-position is highly sensitive to the electronic and steric nature of the chemically modified substituent (55).…”

NAD+ and NAD+ analogues in horse liver alcohol dehydrogenase. Relationship between reactivity and conformation simulated with molecular mechanics

“…Of special interest are those which are modified in the reactive part, the nicotinamide moiety. Kinetic data with such analogues have been published on various dehydrogenases [1][2][3][4][5][6][7][8][9][10][11], but no systematic explanation has so far been provided for their reactivities. In the present paper a qualitative explanation using molecular mechanics calculations is reported.…”