Phosphorylases amino acids

Be Pyridoxine, pyridoxal, pyridoxamine Coenzyme in transamination and decarboxylation of amino acids and glycogen phosphorylase role in steroid hormone action Disorders of amino acid metabolism, convulsions... 

Six compounds have vitamin Bg activity (Figure 45-12) pyridoxine, pyridoxal, pyridoxamine, and their b -phosphates. The active coenzyme is pyridoxal 5 -phos-phate. Approximately 80% of the body s total vitamin Bg is present as pyridoxal phosphate in muscle, mostly associated with glycogen phosphorylase. This is not available in Bg deficiency but is released in starvation, when glycogen reserves become depleted, and is then available, especially in liver and kidney, to meet increased requirement for gluconeogenesis from amino acids. 

Pyridoxal phosphate is a coenzyme for many enzymes involved in amino acid metabolism, especially in transamination and decarboxylation. It is also the cofactor of glycogen phosphorylase, where the phosphate group is catalytically important. In addition, vitamin Bg is important in steroid hormone action where it removes the hormone-receptor complex from DNA binding, terminating the action of the hormones. In vitamin Bg deficiency, this results in increased sensitivity to the actions of low concentrations of estrogens, androgens, cortisol, and vitamin D. 

Pantothenic acid is present in coenzyme A and acyl carrier protein, which act as carriers for acyl groups in metabolic reactions. Pyridoxine, as pyridoxal phosphate, is the coenzyme for several enzymes of amino acid metabolism, including the aminotransferases, and of glycogen phosphorylase. Biotin is the coenzyme for several carboxylase enzymes. 

The enzyme has been isolated from both eukaryotic and prokaryotic organisms [2] and functions in the purine salvage pathway [1,3]. Purine nucleoside phosphorylase isolated from human erythrocytes is specific for the 6-oxypurines and many of their analogs [4] while PNPs from other organisms vary in their specificity [5]. The human enzyme is a trimer with identical subunits and a total molecular mass of about 97,000 daltons [6,7]. Each subunit contains 289 amino acid residues. 

In the analysis of the structural data of other protein kinases, it is noted that only cAPK has been crystallized with its specific peptide inhibitor. Nevertheless, three other structures of protein kinases compared with the structure of the cAPK-PKI complex provide substantial evidence for the conservation of the substrate binding cleft. The substrate binding cleft of the phosphorylase kinase structure has been analyzed in detail and it is clear that all amino acids of the known specific substrate can be built into the PKI model and all required corresponding charges can be found in the cleft of the phosphorylase kinase structure. In the CK-1 structure determined without a peptide, the requirement of the peptide specificity resides on the P-3 site, which has to be phosphorylated. An analysis of the surface charges of the cleft of the CK-1 structure reveals the exact correspondence of the residues required to interact with a phosphorylated substrate at this site. 

Pyridoxal phosphate is an essential cofactor in the glycogen phosphorylase reaction its phosphate group acts as a general acid catalyst, promoting attack by Pj on the glycosidic bond. (This is an unusual role for this cofactor its more typical role is as a cofactor in amino acid metabolism see Fig. 18-6.)... 

The control of glycogen phosphorylase by the phosphorylation-dephosphorylation cycle was discovered in 1955 by Edmond Fischer and Edwin Krebs50 and was at first regarded as peculiar to glycogen breakdown. However, it is now abundantly clear that similar reactions control most aspects of metabolism.51 Phosphorylation of proteins is involved in control of carbohydrate, lipid, and amino acid metabolism in control of muscular contraction, regulation of photosynthesis in plants,52 transcription of genes,51 protein syntheses,53 and cell division and in mediating most effects of hormones. 

Polynucleotide phosphorylase was used to produce RNA of random sequence, the composition of which reflected the mixture of nucleoside diphosphates in the reaction mixture. Mixed polynucleotides containing two bases were used in the incorporating system and shown to incorporate a pattern of amino acids consistent with a triplet code, but the observed incorporation could not define the code sequence. 

It is of interest to compare the tertiary structure of AspAT with that of other PLP-dependent enzymes. Some PLP enzymes whose primary structures are quite different from AspAT exhibit similar tertiary structures. Such enzymes are a>-amino acid pyruvate aminotransferase,341 phosphoserine aminotransferase351 and tyrosine-phenol lyase361 (Phillips, R., personal communication). Similarity in tertiary structure among these PLP enzymes may lead to the idea that many PLP-dependent enzymes share the same ancestor protein. There are PLP enzymes belonging to its own category, such as glycogen phosphorylase and tryptophan synthase.37 381 These enzymes do not share any similarities in either primary or tertiary structures with AspAT. 

The polypeptide of type-L isozyme is composed of 916 amino acid residues. There are two covalently modified amino adds about one-fourth of the amino-terminal threonines are blocked by an acetyl group, and a lysyl residue (Lys762) is linked to the cofactor pyridoxal-P. The partial amino-terminal acetylation appeared to be a natural feature of type-L isozyme, and both the blocked and unblocked forms of type-L phosphorylase may exist in potato tubers. 

The transit peptide of the type-L phosphorylase isozyme of potato amyloplast has the following amino acid composition and sequence ... 

Another interesting feature of the transit peptide of the type-L phosphorylase isozyme is the high content of histidine the peptide contains five histidyl residues in a total of 50 amino acid residues, in contrast to other transit peptides that contain no or little, if any, histidyl residues. It is not known if these residues have any special role other than increasing the positive charge of the peptide. 

Amino acid sequence comparison of a-glucan phosphorylases. H, the potato type-H isozyme L, the potato type-L isozyme and R, rabbit muscle phosphotylase.791 The type-H isozyme sequence is used as the reference sequence only the amino acid residues that are nonidentical in the type-L isozyme and rabbit muscle enzyme are indicated. (From Biol. Chem., 266 (28), 18453 (1991)). 

Previously, it has been shown that most of the residues directly interacting with AMP as well as the phosphorylatable Ser14 and its surroundings in the rabbit muscle enzyme are far less conserved in the potato type-L isozyme sequence.63 Likewise, the amino-terminal region of the potato type-H isozyme is completely different from that of the rabbit muscle enzyme over the first 80 amino acid residues, in which the sites of covalent phosphorylation and of allosteric regulation by AMP are all included. These variances in sequence are compatible with the lack of regulation in the plant phosphorylase isozymes. 

Yanase, M., Takata, H., Fujii, K., Takaha, T., and Kuriki, T. 2005. Cumulative effect of amino acid replacements results in enhanced thermostability of potato-type L a-glucan phosphorylase. Appl. Environ. Microbiol., 71, 5433-5439. 

Pyridoxamine amino acids and glycogen phosphorylase modulation of steroid hormone action ... 

Vitamin Be has a central role in the metabolism of amino acids in transaminase reactions (and hence the interconversion and catabolism of amino acids and the synthesis of nonessential amino acids), in decarboxylation to yield biologically active amines, and in a variety of elimination and replacement reactions. It is also the cofactor for glycogen phosphorylase and a variety of other enzymes. In addition, pyridoxal phosphate, the metabolically active vitamer, has a role in the modulation of steroid hormone action and the regulation of gene expression. 

Muscle pyridoxal phosphate is released into the circulation (as pyridoxal) in starvation as muscle glycogen reserves are exhausted and there is less requirement for glycogen phosphorylase activity. Under these conditions, it is potentially available for redistribution to other tissues, especially the liver and kidneys, to meet the increased requirement for gluconeogenesis from amino acids (Black et al., 1978). However, during both starvation and prolonged bed rest, there is a considerable increase in urinary excretion of 4-pyridoxic acid, suggesting that much of the vitamin Be released as a result of depletion of muscle glycogen and atrophy of muscle is not redistributed, but rather is ca-tabolized and excreted (Cobum et al., 1995). 

Unlike other pyridoxal phosphate-dependent enzymes, in which it is the carbonyl group that is essential for catalysis, the internal Schiff base between pyridoxal phosphate and lysine in glycogen phosphorylase can be reduced with sodium borohydride without affecting catalytic activity. Thus, while pyridoxal phosphate is essential for phosphorylase activity, it does not act by the same kind of mechanism as in amino acid metabolism. 

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