Synthesis of coenzymically active soluble and insoluble macromolecularized NAD+ derivatives.
Alkylation at N-1 of the NAD+ adenine ring with 3,4-epoxybutanoic acid, followed by chemical reduction to the alkali-stable NADH form and alkaline Dimroth rearrangement, gave the NADH derivative alkylated at the exocyclic adenine amino group. Enzymic reoxidation of the latter derivative gave nicotinamide-6-(2-hydroxy-3-carboxypropylamino)purine dinucleotide, a functionalized NAD+ analogue carrying an omega-carboxyalkyl side-chain at the exocyclic adenine amino group. Carbodiimide coupling of the latter derivative to high-molecular-weight water-soluble (polyethyleneimine, polylysine) and insoluble (aminohexyl-Sepharose) polymers gave the corresponding macromolecularized NAD+ analogues. These derivatives have been shown to be enzymically reducible. The polyethyleneimine and polylysine analogues showed a substantial degree of efficiency relative to free NAD+ with rabbit muscle lactate dehydrogenase (60 and 25% respectively) but a lower one with yeast alcohol dehydrogenase and Bacillus subtilis alanine dehydrogenase (2-7%). The polyethyleneimine derivative entrapped in cellulose triacetate fibres together with the lactate dehydrogenase was operationally stable during repetitive use.
D000409:Alanine; D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000594:Amino Acid Oxidoreductases / Q000378:metabolism; D000818:Animals; D001412:Bacillus subtilis / Q000201:enzymology; D002849:Chromatography, Gas; D002850:Chromatography, Gel; D002855:Chromatography, Thin Layer; D004590:Electrophoresis, Paper; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008239:Lysine; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008722:Methods; D009132:Muscles / Q000201:enzymology; D009243:NAD / Q000031:analogs & derivatives*; D009904:Optical Rotation; D010455:Peptides; D011108:Polymers; D011817:Rabbits; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D012995:Solubility; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
D010455:Peptides; D011108:Polymers; D009243:NAD; D000429:Alcohol Oxidoreductases; D007770:L-Lactate Dehydrogenase; D000594:Amino Acid Oxidoreductases; D008239:Lysine; D000409:Alanine