Glucose biotin

FIGURE 5.25. Avidin-biotin construction of a monolayer glucose oxidase electrode with an attached ferrocenium cosubstrate and cyclic voltammetric response in a phosphate buffer (pH 8) at 25°C and a scan rate of 0.04 V/s. a attached ferrocene alone, h In the presence of 0.5 M glucose, c Variation of the inverse of the plateau current with the inverse of substrate concentration. Adapted from Figure 1 in reference 24, with permission from the American Chemical Society. 

Fixation of carbon dioxide by biotin-enzyme complexes is not unique to acetyl-CoA, and another important example occurs in the generation of oxaloacetate from pyravate in the synthesis of glucose from non-carbohydrate sources (gluconeogene-sis). This reaction also allows replenishment of Krebs... 

In the ruminant mammary tissue, it appears that acetate and /3-hydroxybutyrate contribute almost equally as primers for fatty acid synthesis (Palmquist et al. 1969 Smith and McCarthy 1969 Luick and Kameoka 1966). In nonruminant mammary tissue there is a preference for butyryl-CoA over acetyl-CoA as a primer. This preference increases with the length of the fatty acid being synthesized (Lin and Kumar 1972 Smith and Abraham 1971). The primary source of carbons for elongation is malonyl-CoA synthesized from acetate. The acetate is derived from blood acetate or from catabolism of glucose and is activated to acetyl-CoA by the action of acetyl-CoA synthetase and then converted to malonyl-CoA via the action of acetyl-CoA carboxylase (Moore and Christie, 1978). Acetyl-CoA carboxylase requires biotin to function. While this pathway is the primary source of carbons for synthesis of fatty acids, there also appears to be a nonbiotin pathway for synthesis of fatty acids C4, C6, and C8 in ruminant mammary-tissue (Kumar et al. 1965 McCarthy and Smith 1972). This nonmalonyl pathway for short chain fatty acid synthesis may be a reversal of the /3-oxidation pathway (Lin and Kumar 1972). 

To replace losses, oxaloacetate can be synthesized from pyruvate and C02 in a reaction that uses ATP as an energy source. This is indicated by the heavy gray line leading downward to the right from pyruvate in Fig. 10-1 and at the top center of Fig. 10-6. This reaction depends upon yet another coenzyme, a bound form of the vitamin biotin. Pyruvate is formed from breakdown of carbohydrates such as glucose, and the need for oxaloacetate in the citric acid cycle makes the oxidation of fats in the human body dependent on the concurrent metabolism of carbohydrates. 

Cholera toxin B subunit-biotin labeled (lyophilized powder, biotin content 0.9mol/mol protein), peroxidase-labeled IgG anti-rabbit antibody (HRP-Ab, from goat, protein content 0.8mg/ml, affinity isolated antibody), anti-cholera toxin (from rabbit, protein content 48mg/ml, purified toxin from Vibrio cholerae), biotin monoclonal anti-rabbit IgG -y-chain specific (from mouse, protein content 4.2mg/ml), glucose oxidase-biotinamidocaproyl labeled (GOX-B, from Aspergillus niger, lyophilized powder containing 40-70% protein, 137 U/mg), polyoxyeth-ylene-sorbitan monolaurate (Tween 20), bovine serum albumin (fraction... 

Fig. 26.2. Amperometric immunosensors set-up using a biotinylated copolymer poly(pyrrole-biotin, pyrrole-lactitob-ionamide) coated platinum or glassy carbon electrodes and three enzymatic markers (GOX-B, PPO-B, HRP-Ab) for the detection of cholera antitoxin. (A) HRP-immunosensor, (B) GOX-B-immunosensor, (C) PPO-B-immunosensor. Mred/Mox = hydroquinone/quinone Gox = biotinylated glucose oxidase PPO — biotinylated polyphenol oxidase HRP-Ab = peroxidase-labeled IgG anti-rabbit antibody.

Claisen reactions involving acetyl-CoA are made even more favourable by first converting acetyl-CoA into malonyl-CoA by a carboxylation reaction with CO2 using ATP and the coenzyme biotin (Figure 2.9). ATP and CO2 (as bicarbonate, HC03-) form the mixed anhydride, which car-boxy lates the coenzyme in a biotin-enzyme complex. Fixation of carbon dioxide by biotin-enzyme complexes is not unique to acetyl-CoA, and another important example occurs in the generation of oxaloacetate from pyruvate in the synthesis of glucose from non-carbohydrate sources... 

Because radionuclides used as markers have many drawbacks, they are more and more often replaced by enzymatic markers (horseradish peroxidase, alkaline phosphatase, glucose oxidase, [3-galactosidase). In order to enhance the sensitivity of determinations, fluorescent markers have been introduced. Another solution relies on avidin—biotin bonds. 

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