Thiamine pyrophosphokinase

Biosynthesis of pyrophosphate (5) from pyrimidine phosphate (47) and thia2ole phosphate (48) depends on the activity of five en2ymes, four of them kinases (87). In yeasts and many other organisms, including humans, pyrophosphate (5) can be obtained from exogenous thiamine in a single step cataly2ed by thiamine pyrophosphokinase (88). 

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... 

Genetic defects of the tissue thiamin transport protein and thiamin pyrophosphokinase cause megaloblastic anemia, presumahly as a result of impaired synthesis of pentoses for DNA synthesis from low activity of trans-ketolase (Section 6.3.2). In many cases, this megaloblastic anemia is thiamin-responsive, suggesting that the defect must be because of low af nity of either the transport protein or thiarnin pyrophosphokinase for its substrate (Neufeld et al., 2001). 

Thiamine uptaken into the cell is phosphorylated to TDP by the enzyme thiamine pyrophosphokinase. TDP is then further phosphorylated to thiamine triphosphate (TTP) or is dephosphorylated to thiamine monophosphate (TMP). 

Fig. 2 Intercellular trafficking and thiamine and thiamine esters in brain. TMP thiamine monophosphate, TDP thiamine diphosphate, TTP thiamine triphosphate, TPKinase thiamine pyrophosphokinase...

Thiamine is absorbed by a pathway that is saturable at concentrations of 0.5-1.0 jumol/L. Oral doses in excess of 10 mg do not significantly increase blood or urine concentrations of vitamin Bi. In the human, absorption occurs predominantly in the jejunum and ileum. Some ferns, shellfish, fish, and species of bacteria contain thiami-nase, which cleaves the pyrimidine ring from the thiazole ring. This enzyme causes thiamine deficiency in cattle. In plasma, thiamine is transported bound to albumin and, to a small extent, other proteins. TPP is synthesized in the liver by thiamine pyrophosphokinase. 

Vitamin Bi is an essential co-factor for several enzymes of carbohydrate metabolism such as transketolase, pyruvate dehydrogenase (PDH), pyruvate decarboxylase and a-ketoglutarate dehydrogenase. To become the active co-factor thiamin pyrophosphate (TPP), thiamin has to be salvaged by thiamin pyrophosphokinase or synthesized de novo. In Escherichia coli and Saccharomyces cerevisiae thiamin biosynthesis proceeds via two branches that have to be combined. In the pyrimidine branch, 4-amino-5-hydroxymethy-2-methylpyrimidine (PIMP) is phosphorylated to 4-amino-2-methyl-5-hydroxymethyl pyrimidine diphosphate (PIMP-PP) by the enzyme HMP/HMP-P kinase (ThiD) however, the step can also be catalyzed by pyridoxine kinase (PdxK), an enzyme also responsible for the activation of vitamin B6 (see below). The second precursor of thiamin biosynthesis, 5-(2-hydroxyethyl)-4-methylthiazole (THZ), is activated by THZ kinase (ThiM) to 4-methyl-5-(2-phosphoethyl)-thiazole (THZ-P), and then the thia-zole and pyrimidine moieties, HMP-PP and THZ-P, are combined to form thiamin phosphate (ThiP) by thiamin phosphate synthase (ThiE). The final step, pyrophosphorylation, yields TPP and is carried out by thiamin pyrophosphorylase (TPK). 

Melnick, J. et al (2004) Identification of the two missing bacterial genes involved in thiamine salvage thiamine pyrophosphokinase and thiamine kinase. J. Bacterial, 186 (11), 3660-3662. 

The coupling reaction between THZ-P and HMP-PP, mediated by ThiE, generates ThMP. The latter is converted to ThDP, the active form of the cofactor, by a thiamin phosphate kinase (ThiL) (Figure 5.92). ThDP can also be formed in one step from thiamin using thiamin pyrophosphokinase. In bacteria, this enzyme is called ThiN. In higher organisms, the thiamin pyrophosphokinase is named THI80. 

Thiamine monophosphate is hydrolyzed to thiamine and phosphate. Thiamine diphosphate is formed from thiamine and ATP by thiamine pyrophosphokinase in microorganisms, plants and animals. 

Thiamin phosphate derivatives can be readily interconverted (Figure 5.5), but the enzymes involved in these reactions remain poorly charaeterized. Only two of them, the essential thiamin pyrophosphokinase and the 25 kDa thiamin triphosphatase, have been charaeterized at the molecular level. 

Ajjjawi, I., Rodriguez Milla, M.A., Cushman, J., and Shintani, D.K., 2007. Thiamin pyrophosphokinase is required for thiamin cofactor activation in Arabidopsis. Plant Molecular Biology. 65 151 162. 

Thiamine monophosphate (18) Affinity chromatographic isolation of thiamine pyrophosphokinase 253... 

Thiamine is metabolized to TPP by thiamine pyrophosphokinase (EC 2.1.62) in animal cells including red and white blood cells. This enzyme is also present in plants, yeast, and a bacterium (Paracoccus denitrificans) (7). However, in some bacteria, for example in Escherichia coli, thiamine is metabolized to TPP by a two-step reaction catalyzed by thiamine kinase (EC 2.7.1.89) and TMP kinase (EC 2.7.4.10). Thiamine pyrophosphate is further metabolized to TI P in yeast, animal tissues, and human red blood cells. Evidence has been obtained which indicates that cytosolic adenylate kinase (EC 2.7.4.3) catalyzes TIP synthesis from TPP in vitro (8) and in vivo (3). The enzyme system involved in thiamine metabolism to TTP in human red blood cells was recently identified, purified, and reconstituted (9). 

H Sanemori, T Kawasaki. Purification and properties of thiamine pyrophosphokinase... 

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