Separation transaminase

PEP-CK version of C4 metabolism. This uses just one ATP per carbon fixation. The little cycle at the top is actually a cheat two separate transaminase reactions occur regeneration of glutamate in the mesophyll and ketoglutarate in the bundle sheath requires pyruvate to be transaminated to alanine by glutamate in the bundle sheath, and shunted out to be transaminated back to pyruvate by ketoglutarate in the mesophyll. 

Amino Acid Amide Transaminases Meister and co-workers (18, 248, 260, 270, 271, 325, 354-356) have established the existence of two separate transaminases which catalyze the following reactions ... 

Demonstrating the existence of multiple transaminases in the late 1930s and early 1940s was very difficult. Chromatographic separation of amino acids was not available until the mid-1940s and paper chromatography was not in common use until 1948-1950 when Hird and Rowsell finally showed the existence of the wide range of transaminases and the universality of the transamination process. 

Serum Preparation Whole blood was collected after decapitation of the animal and allowed to clot at room temperature for 30 min. The serum was then separated from the clot by centrifugation at 5000 rpm at U C. Aliquots of the serum were immediately taken and refrigerated for the determination of glutamic-pyruvic transaminase and lactic dehydrogenase. An additional 0.5 ml of serum from each animal was treated with a few milligrams of sodium fluoride and refrigerated for later glucose determinations. The rest of the serum was frozen for the remainder of the clinical chemistry determinations. 

N4. Nisselbaum, J. S., and Bodansky, O., Immunochemical and kinetic properties of anionic and cationic glutamic-oxaloacetic transaminases separated from human heart and human liver. J. Biol. Chem. 239, 4232-4236 (1964). 

The X-ray induced mutant C-2A was cultured and harvested as described earlier (10). DOVA-transaminase was purified as demonstrated in (11). The preparation of tRNA-fractions and the corresponding ligase preparation as well as the run-off fraction, containing the transaminases, ALA-dehydratase and porphobilinogenase were performed following (12). G-1-SA-pyrrole was formed by the condensation of the compound with ethylacetoacetate as described in (13). Protochlorophyllide (Pchlide) was isolated following (14). The separation into divinyl- and monovinyl-Pchlide was performed as described in (15). 

The toxic effects of synthetic polymers include inhibition of hepatic microsomal oxidase enzymes, stimulation of liver transaminases, hepatosplenomegaly, thymic involution, anaemia, and decrease in bone marrow cells Fortunately, many neutral synthetic polymers do not show toxic effects. For those polymers which exhibit toxic effects, e.g. polyanions, it is sometimes possible to separate the toxicity from antitumour, antiviral and immunological effects In general the toxicity of polyanions increases with molecular weight (particularly for M > 50,000). 

Fig. 12.41. Differentiation of fresh liver (1) from frozen and thawed liver (2) by electrophoretic separation of glutamate-oxalacetate transaminases (a) GOT sarcoplasm, (b) GOT mitochondria (according to Hamm 2indMasic, 1975)...

Such membrane damage has been observed by freezing and thawing of tissue, for example, of muscle tissue, in which the isoenzymes of glutamate oxalacetate transaminase (GOT) bound to mitochondrial membranes are partially released and found in the sarcoplasm. The pressed sap collected from fresh unfroze meat has only sarcoplasm enzymes, while the frozen and thawed meat has, in addition, the isoenzymes derived from mitochondria. The GOT isoenzymes can be separated by electrophoresis (Fig. 12.41). This procedure is also apphcable to fish. 

Rudman and Meister IJ ) first showed the presence of a transaminase in cell-free extracts of E. colt that catalyze transamination reactions between glutamate and isoleucine, valine, leucine, norleucine, and norvaline. These monocarboxylic amino acids transaminated with each other as well as with glutamine. Preparations of an E. cdi mutant which did not respond to a-keto- 8-methylvalerate was unable to transaminate isoleucine or valine. The transaminase responsible for activity with the branched-chain amino acids was separated from other transaminases and considerably purified by standard methods of protein purification. It was shown to... 

The recombinant (R)-transaminase prepared from cell extracts of rec E. coli SC16577 (40 g wet cells/200 ml suspension) was suspended in buffer containing 50 mM potassium phosphate pH 7,1 mM dithiothreitol, and 10 xM pyridoxal phosphate using a microfluidizer at 12000 psi followed by separation of cell debris. The reaction mixture contained 10 mL of this cell extract, keto acid 90 (6.0 g), D,L-alanine (12.0 g), pyridoxal phosphate (2.6 mg), dithiothreitol (15.0 mg) and 0.1 M potassium phosphate buffer pH 7.5 (80 mL). After incubating at 30°C with gentle shaking at 50 rpm for 39 h, the reaction was complete with 95% in-process yield and the product (R)-88a was isolated in 79% yield (5.4g) with >99% ee [155]. 

Synthesis of 2-amino-1,3,4-butanetriol by a transketolase/ transaminase enzyme cascade via continuous flow dual reactor. The enzymes were immobilized inside the silica microreactors separately. 

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