Thiamin-binding Proteins

HMP, forming first HMP-P (8, salvage activity) and then HMP-PP (9, biosynthetic activity)/ Structural studies have provided an explanation for this novel dual kinase activity." The ABC transporter involved in thiamin uptake has also been identified and the periplasmic thiamin-binding protein has been structurally characterized. ... 

A very good separation of thiamine-binding protein from various rat organs was achieved on a TSK-gel G 3000SW column [109]. The method proved to be rapid and sensitive. 

Affinity chromatographic purification of kallikrein from human serum 378 Affinity chromatographic isolation of thiamine-binding protein from 379 chicken egg-white... 

Thiamin transport has been nearly exclusively studied in mammalian cells. However, a few studies carried out in bacteria show that thiamin can be transported via ABC transporters. These transporters comprise a periplasmic thiamin binding protein in gram-negative bacteria such as E. coli and Salmonella typhimurium. These proteins are generally not specific for thiamin as they also bind ThMP and ThDP. Examination of the three-dimensional structure of an E. coli thiamin binding protein revealed structural similarities with thiami-nase I, suggesting a common ancestor (Soriano et al. 2008). 

Thiamin-binding proteins have been described in many animal and plant tissues, but none has been characterized at the molecular level. They exist in eggs... 

Soriano, E.V., Rajashankar, K.R., Hanes, J.W., Bale, S., Begley, T.P., and Ealick, S.E., 2008. Structural similarities between thiamin-binding protein and thiaminase-1 suggest a common ancestor. Biochemistry. 47 1346-1357. 

Thiamine binds to an enzyme before the enzyme is activated. The enzyme also binds to the substrate (a large protein). Without the coenzyme thiamine, no chemical reaction would occur. The coenzyme is the chemical reagent. The protein molecule (the enzyme) helps and mediates the reaction by controlling stereochemical, energetic, and entropic factors, but it is nonessential to the overall course of reactions that it catalyzes. A special name, vitamins, is given to coenzymes that are essential to the nutrition of the organism. 

A peculiarity of thiamine is that the vitamin can easily become inactivated. An early instance was seen in 1941 when commercially reared mink became paralyzed (Chastek paralysis), a disorder which could be cured by giving the animals thiamine. The problem was traced to their having been fed fish that had partially decomposed. Later work showed that in decayed fish a microbial enzyme had been released, thiaminase, which destroyed the thiamine normally present in the food. A rather different process occurs when horses or cows are allowed to graze on bracken. This contains a protein which binds to thiamine, so reducing its availability. Once again the condition can be treated by administering the vitamin. 

Thiamine pyrophosphate (TPP) (37), a derivative of vitamin Bi (38), contains two substituted heterocycles, the pyrimidine and thiazolium rings, connected by a methylene bridge. The reactive portion of this coenzyme is the thiazolium ring, the pyrimidine portion (as well as the pyrophosphate group) being important in binding interactions with proteins... 

The ALS isolated as described in Table III displayed typical Michaelis-Menten kinetics with respect to pyruvate with a Km of 2.44 mM. Substrate concentrations as high as 50x Km had no effect on the rate of the reaction. Thiamine pyrophosphate, FAD and Mg(2+) were an absolute requirement for catalysis by the purified enzyme. These properties are consistent with observations made by others (30). Optimum activity was obtained at pH 7.1 and 37C, which were also the best conditions for inhibition by TP. There was no significant difference in the 1(50) value of TP whether ALS was taken after step 2 or 5, indicating low potential for non-specific binding of the herbicide to other proteins. 

Nilsson U, Meshalkma L, Lindqvist Y, Schneider G (1997) Examination of substrate binding in thiamin diphosphate-dependent transketolase by protein crystallography and site-directed mutagenesis. I Biol Chem 272 1864— 1869... 

However, another study was an example of nature appreciation—the structure of thiamine was varied to learn what was special about the particular thiazolium derivative that was natural thiamine (11). As a chemical catalyst—ignoring the question of what effect changes would have on the ability of the coenzyme to bind to the proteins that have evolved to use it—thiamine proved to be the optimal relative to other related structures because of a balance of catalytic ability and chemical stability. The anion 4 derived from an imidazolium ring instead of a thiazolium ring was a weaker catalyst but was more stable in water (10). [The imidazolium aiuon and its dihydro derivative have proven to be very useful metal ion ligands, including... 

Ethanol is an "antivitamin" that decreases the cellular content of almost every coenzyme. For example, ethanol inhibits the absorption of thiamine, and acetaldehyde produced from ethanol oxidation displaces pyridoxal phosphate from its protein binding sites, thereby accelerating its degradation. 

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