Thiamin Transport

Most of the vitamins and cofactors discussed in previous chapters of the text would be needed during starvation because many of the essential metabolic pathways must continue to operate. Among the most obvious vitamins needed for those pathways are pyri-doxal phosphate (for the transamination of amino acids), niacin and riboflavin (for electron transport), thiamin (for the oxidative decarboxylation of pyruvate, a-ketoglu-tarate, and the branched-chain amino acids), biotin (for the carboxylation of pyruvate), and cobalamin (for the conversion of methylmalonyl Co A to succinyl Co A). 

Another thiamin transporter, ThTR2, is the product of the SLC19A3 gene. It transports thiamin with a very high apparent affinity = 20-100 nM). 

A.88.3.2 Thiamin transporter (Thiamin ECF transporter S component) ThiT L. lactis Majsnerowska et al. 2013... 

In humans, thiamine is both actively and passively absorbed to a limited level in the intestines, is transported as the free vitamin, is then taken up in actively metabolizing tissues, and is converted to the phosphate esters via ubiquitous thiamine kinases. During thiamine deficiency all tissues stores are readily mobilhed. Because depletion of thiamine levels in erythrocytes parallels that of other tissues, erythrocyte thiamine levels ate used to quantitate severity of the deficiency. As deficiency progresses, thiamine becomes indetectable in the urine, the primary excretory route for this vitamin and its metaboHtes. Six major metaboHtes, of more than 20 total, have been characterized from human urine, including thiamine fragments (7,8), and the corresponding carboxyHc acids (1,37,38). 

SMVT Sodium-dependent multivitamin transporter SVCT Sodium-dependent vitamin C transporter THTR High affinity thiamine transporter... 

Organic cation transporters OCT1/2 tetraethylammonium (TEA), thiamine,... 

The water-soluble vitamins generally function as cofactors for metabolism enzymes such as those involved in the production of energy from carbohydrates and fats. Their members consist of vitamin C and vitamin B complex which include thiamine, riboflavin (vitamin B2), nicotinic acid, pyridoxine, pantothenic acid, folic acid, cobalamin (vitamin B12), inositol, and biotin. A number of recent publications have demonstrated that vitamin carriers can transport various types of water-soluble vitamins, but the carrier-mediated systems seem negligible for the membrane transport of fat-soluble vitamins such as vitamin A, D, E, and K. 

Rajgopal, A., et al. SLC19A3 encodes a second thiamine transporter ThTr2. Biochim. Biophys. Acta 2001, 3537, 175-178. 

Dutta, B., et al. Cloning of the human thiamine transporter, a member of the folate transporter family. J. Biol. Chem. 1999, 274, 31925-31929. 

The BBB also has sodium- and pH-independent transporters of organic cations. They are important for the homeostasis of choline and thiamine in the brain and for the permeation of cationic drugs like propranolol, lidocaine, fentanyl, Hl-an-... 

In brain, as in most mammalian cells, thiamine occurs predominantly in the form of TDP, the remainder being made up of thiamine monophosphate (10%), thiamine triphosphate (5-10%) and trace amounts of free thiamine. Thiamine is transported into brain and phosphory-lated by the action of thiamine pyrophosphokinase, and inhibition of this enzyme by thiamine antagonists such as pyrithiamine results in decrease synthesis of TDP. Treatment of experimental animals with pyrithiamine results in a generalized reduction of TDP concentrations and an early selective loss in activity of a-KGDH in regions... 

Most coenzymes have aromatic heterocycles as major constituents. While enzymes possess purely protein structures, coenzymes incorporate non-amino acid moieties, most of them aromatic nitrogen het-erocycles. Coenzymes are essential for the redox biochemical transformations, e.g., nicotinamide adenine dinucleotide (NAD, 13) and flavin adenine dinucleotide (FAD, 14) (Scheme 5). Both are hydrogen transporters through their tautomeric forms that allow hydrogen uptake at the termini of the quinon-oid chain. Thiamine pyrophosphate (15) is a coenzyme that assists the decarboxylation of pyruvic acid, a very important biologic reaction (Scheme 6). 

Fig. 3. Intracellular transport route of the SFV spike glycoproteins from the endoplasmic reticulum (ER), over the Golgi apparatus, to the plasma membrane (PM). The cis cisternae do not react positively for add phosphatase or thiamin pyrophosphatase, and do not label with ridn in thin frozen sections. The medial cisternae do not react positively for thiamin pyrophosphatase or acid phosphatase, but label with ridn. The trans cisternae are positive for all of these markers.

In this reaction, pyruvic acid is oxidized to carbon dioxide with formation of acetyl-SCoA and NAD+ is reduced to NADH. As noted in chapter 15, this reaction requires the participation of thiamine pyrophosphate as coenzyme. Here too the NADH formed is converted back to NAD+ by the electron transport chain. As noted above, the acetyl-SCoA is consumed by the citric acid cycle and CoASH is regenerated. 

The transport of amino acids at the BBB differs depending on their chemical class and the dual function of some amino acids as nutrients and neurotransmitters. Essential large neutral amino acids are shuttled into the brain by facilitated transport via the large neutral amino acid transporter (LAT) system [29] and display rapid equilibration between plasma and brain concentrations on a minute time scale. The LAT-system at the BBB shows a much lower Km for its substrates compared to the analogous L-system of peripheral tissues and its mRNA is highly expressed in brain endothelial cells (100-fold abundance compared to other tissues). Cationic amino acids are taken up into the brain by a different facilitative transporter, designated as the y system, which is present on the luminal and abluminal endothelial membrane. In contrast, active Na -dependent transporters for small neutral amino acids (A-system ASC-system) and cationic amino acids (B° system), appear to be confined to the abluminal surface and may be involved in removal of amino acids from brain extracellular fluid [30]. Carrier-mediated BBB transport includes monocarboxylic acids (pyruvate), amines (choline), nucleosides (adenosine), purine bases (adenine), panthotenate, thiamine, and thyroid hormones (T3), with a representative substrate given in parentheses [31]. 

During the first 3 h after Intravenous injection of 1 that followed administration of thiamine, urinary excretion of the oxime was about 12.7% below that during the corresponding period of the control experiment during the remainder of the run, it was 62.2% above that during the same period of the control experiment. Inasmuch as intravenous injection of 900 mg of sodium jg-aminohippurate with I decreased by only 6.3% the urinary excretion of I during the first 3 h after its administration, the tubular transport mechanisms for 1 and for jg-amino-hippurate probably are different. 

Non-enzymatic proteins found in both parasites and humans, but exhibiting different pharmacological profiles between the two species (e.g., thiamine transporter)... 

Amprolium (Fig. 5.7) is a vitamin IT analogue. It is a competitive antagonist of the thiamine transport mechanism. Amprolium has been used as a coccidi-ostat mainly in chickens, laying hens, turkeys, and ruminants. It is available as a soluble powder for addition to drinking water (60-240 mg/L) or as a premix, usually in combination with ethopabate and/or sulfaquinoxaline, for mixing with the feed (125-500 mg/kg feed). A withdrawal period of 3 days is required for chickens. 

Vitamins are chemically unrelated organic compounds that cannot be synthesized by humans and, therefore, must must be supplied by the diet. Nine vitamins (folic acid, cobalamin, ascorbic acid, pyridoxine, thiamine, niacin, riboflavin, biotin, and pantothenic acid) are classified as water-soluble, whereas four vitamins (vitamins A, D, K, and E) are termed fat-soluble (Figure 28.1). Vitamins are required to perform specific cellular functions, for example, many of the water-soluble vitamins are precursors of coenzymes for the enzymes of intermediary metabolism. In contrast to the water-soluble vitamins, only one fat soluble vitamin (vitamin K) has a coenzyme function. These vitamins are released, absorbed, and transported with the fat of the diet. They are not readily excreted in the urine, and significant quantities are stored in Die liver and adipose tissue. In fact, consumption of vitamins A and D in exoess of the recommended dietary allowances can lead to accumulation of toxic quantities of these compounds. 

Pyruvate oxidase. The soluble flavoprotein pyruvate oxidase, which was discussed briefly in Chapter 14 (Fig. 14-2, Eq. 14-22), acts together with a membrane-bound electron transport system to convert pyruvate to acetyl phosphate and C02.319 Thiamin diphosphate is needed by this enzyme but lipoic acid is not. The flavin probably dehydrogenates the thiamin-bound intermediate to 2-acetylthiamin as shown in Eq. 15-34. The electron acceptor is the bound FAD and the reaction may occur in two steps as shown with a thiamin diphosphate radical intermediate.3193 Reaction with inorganic phosphate generates the energy storage metabolite acetyl phosphate. 

Carbohydrate metabolism provides the main energy source in coccidia. Diets deficient in thiamin, riboflavin, or nicotinic acid—all cofactors in carbohydrate metabolism—result in suppression of parasitic infestation of chickens by E tenella and E acervulina. A thiamin analog, amprolium—1-[(4-amino-2-propyl-5-pyrimidinyl)-methyl]-2-picolinium chloride—has long been used as an effective anticoccidial agent in chickens and cattle with relatively low host toxicity. The antiparasitic activity of amprolium is reversible by thiamin and is recognized to involve inhibition of thiamin transport in the parasite. Unfortunately, amprolium has a rather narrow spectrum of antiparasitic activity it has poor activity against toxoplasmosis, a closely related parasitic infection. 

Facilitated diffusion involves carrier-mediated transport down a concentration gradient. The existence of the carrier molecules means that diffusion down the concentration gradient is much greater than would be expected on the basis of the physicochemical properties of the drag. A much larger number of substances are believed to be transported by facilitated diffusion than active transport, including vitamins such as thiamine, nicotinic acid, riboflavin and vitamin B6, various sugars and amino acids. 

Pyruvate produced by the glycolytic pathway may be transported into the mitochondria (via an antiport with OH"), where it is converted to acetyl-CoA by the action of the enzyme complex pyruvate dehydrogenase. The pertinent enzyme activities are pyruvate dehydrogenase (PD), lipoic acid acetyltransferase, and dihydrolipoic acid dehydrogenase. In addition, several cofactors are utilized thiamine pyrophosphate (TPP), lipoic acid, NAD+, Co A, and FAD. Only Co A and NAD+ are used in stoichiometric amounts, whereas the others are required in catalytic amounts. Arsenite and Hg2+ are inhibitors of this system. The overall reaction sequence may be represented by Figure 18.5. The NADH generated may enter the oxidative phosphorylation pathway to generate three ATP molecules per NADH molecule reduced. The reaction is practically irreversible its AGq = -9.4 kcal/mol. 

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