Copyright © 2000-2011 Mark Brandt, Ph.D. 36 Vitamins and Coenzymes Vitamins are compounds that are required in the diet, either because the organism cannot synthesize them, or because the rate of usage by the organism typically exceeds the rate of synthesis of the compound. In nearly all cases, only very small amounts of these compounds are required. Vitamins are generally classed as either water-soluble or fat-soluble. The water-soluble vitamins generally act as precursors to coenzymes; the functions of the fat-soluble vitamins are more diverse and less easily categorized. The water-soluble vitamins are readily excreted in the urine; toxicity as a result of overdose is therefore rare. However, with few exceptions, the water-soluble vitamins are not stored in large amounts, and therefore must be continually supplied in the diet. In contrast, the fat-soluble vitamins are less readily excreted, and are deleterious (and possibly lethal) in high doses. Many of the fat-soluble vitamins are stored; for example, most well nourished individuals have a three month supply of vitamin D. Water soluble vitamins The water-soluble vitamins include the B complex vitamins (the actual B vitamins, biotin, and folic acid) and vitamin C. First we will look at three classes of vitamin-derived coenzymes used to carry electrons: the nicotinamide coenzymes, the flavin coenzymes, and ascorbic acid. Vitamin B3 (niacin) Niacin is the name for both nicotinamide and nicotinic acid, either of which can act as a precursor of nicotinamide coenzymes. Niacin is required for the synthesis of two coenzyme molecules: NAD and NADP. Note the phosphate attached to the 2´-position of the lower ribose ring in NADP, which is the only difference between the molecules. OOHOHCH2NOPOOOOOHOHCH2ONNNNNH2POONH2OOOPOOOOHOHCH2NOPOOOOOHCH2ONNNNNH2POONH2ONicotinamide adeninedinucleotide[NAD]Nicotinamide adeninedinucleotide 2´-phosphate[NADP]NNH2ONOHONiacin(nicotinic acid)Vitamin B3Niacin(nicotinamide)Vitamin B3Copyright © 2000-2011 Mark Brandt, Ph.D. 37 Humans can synthesize nicotinamide cofactors from tryptophan. However, the process is somewhat inefficient; synthesis of 1 mg of niacin requires 60 mg of tryptophan. Niacin deficiency therefore is usually the result of a diet deficient in both niacin and tryptophan. However, some diets contain tryptophan or niacin in a biologically unavailable form. In corn, the niacin is poorly absorbed unless the corn is treated with alkali prior to ingestion. In the rural south of the early 20th century, this preparation step was largely ignored; the symptoms of the resulting pellegra (niacin-deficiency), such as sun-sensitivity and dementia, led to the pejorative term “red-neck” for individuals from this region of the US. Pellegra is also observed in high sorghum diets (sorghum contains niacin-synthesis inhibitors) or in some individuals taking isoniazid (isoniazid is an antibiotic used to treat tuberculosis, but also inhibits niacin uptake and synthesis). Nicotinic acid reduces release of free fatty acids from adipose tissue, and has been used to reduce plasma cholesterol (nicotinamide is inactive for this purpose). However, some individuals cannot tolerate the level of nicotinic acid required. Niacin is required for the synthesis of two coenzyme molecules: NAD and NADP. Note the 2´-phosphate attached to the lower ribose ring in NADP, which is the only structural difference between the molecules. NAD and NADP act as soluble electron carriers between proteins. In effect, these compounds are substrates for enzymes involved in oxidation and reduction reactions. NAD is primarily involved in catabolic reactions. NAD accepts electrons during the breakdown of molecules for energy. In contrast, NADPH (the reduced form of NADP) is primarily involved in biosynthetic reactions; it donates electrons required for synthesizing new molecules. In most cells, NAD levels are much higher than NADH levels, while NADPH levels are much higher than those of NADP. The two possible electronic states for the nicotinamide cofactors are shown below: The oxidized forms of both nicotinamide coenzymes can only accept electrons in pairs. The reduced forms of the coenzymes can only donate pairs of electrons. Note the two changes in the ring during the reduction. The addition of the electron NNH2ORH2 electrons+1 protonNNH2ORH H2 electrons+1 protonOxidizedNAD(P)ReducedNAD(P)HNHNONH2IsoniazidCopyright © 2000-2011 Mark Brandt, Ph.D. 38 pair is accomplished by the addition of a hydride ion to the carbon para to the pyridine nitrogen, and results in the loss of the positive charge on the ring. Nicotinic acid was first synthesized chemically in 1867 from nicotine: The name “niacin” was introduced to remove the association with nicotine and tobacco. Alcohol Dehydrogenase An example of the role of NAD in redox chemistry is provided by the oxidoreductase enzyme liver alcohol dehydrogenase. The name of the enzyme includes the tissue of origin and the substrate. The word “dehydrogenase” is an indication of the fact that the enzyme catalyzes an oxidation-reduction reaction. (“Dehydrogenase” means “catalyzes hydrogen removal”.) Alcohol dehydrogenase can catalyze the oxidation of several different alcohols. In each case it uses NAD as the electron acceptor. The active site is thus moderately non-specific for the alcohol, although it is quite specific for NAD compared to NADP. In the absence of substrate, the alcohol dehydrogenase active site is occupied by water molecules. Note the zinc ion, a metal ion cofactor that is required for catalytic activity (alcohol dehydrogenase actually binds two zinc ions, but the other is thought to have an exclusively structural role). The zinc is bound to three enzyme side-chains (two cysteine residues and a histidine residue). Binding of substrate causes a conformational change that excludes water from the active site, and that positions the substrates in preparation for catalysis. When the substrate binds, the zinc ion coordinates (i.e. binds) to the alcohol oxygen. This bond NNCH3HNO3NOHONicotine Nicotinic acidHNNSSZnHOHNHNOHCys46His67Ser48Cys174His51Copyright ©