共查询到20条相似文献,搜索用时 15 毫秒
1.
Effect of glucose-6-P on the catalytic and structural properties of glycogen phosphorylase a 总被引:8,自引:0,他引:8
A monoclonal antibody to porcine beta-lipotropin has been produced which binds to the N-terminal (gamma-lipotropin) portion of the molecule. The antibody can be used to detect beta-lipotropin as well as other beta-endorphin precursors (predominantly a Mr 38 000 polypeptide) using radiobinding assay or the immunoblotting technique. Purification of the peptides can be readily achieved by affinity chromatography using the monoclonal antibody covalently bound to Sepharose 4B. As the antibody recognises the N-terminal part of beta-lipotropin, it can be used to detect and purify beta-lipotropin and other beta-endorphin precursors in the presence of beta-endorphin. 相似文献
2.
3.
4.
C E Alonso A Coviello 《Comparative biochemistry and physiology. A, Comparative physiology》1986,84(2):271-273
The effects of the bile salts, sodium taurocholate (NaTc) and sodium cholate (NaCh), and toad bile gallbladder (bile) on short-circuit current (SCC) across isolated skin, and sodium taurocholate (NaTc) on isolated bladder of Bufo arenarum toads were tested. Sodium taurocholate (NaTc), sodium cholate (NaCh) and toad bile gallbladder (bile) promoted an increase in SCC, when added to the external side. The stimulatory effect was reversible after rinsing the preparation for 60 min. Implications on in vivo renal function of these results are discussed. 相似文献
5.
Rabbit skeletal muscle phosphorylase b was separated into two fractions by column chromatography on AMP-Sepharose. The first fraction protein was eluted by glucose-6-phosphate while the second fraction protein was eluted in an AMP concentration gradient. The bulk of the protein eluate was represented by the first fraction protein. Chromatography of phosphorylase b from bovine skeletal muscle under identical conditions also resulted in two fractions, however, with a reverse correlation: the bulk protein of this fraction was eluted by AMP. It was shown that the two phosphorylase b forms eluted by glucose-6-phosphate and AMP differ by their kinetic and physico-chemical properties as well as by the SH-group reactivity. The phosphorylase b forms eluted by the nucleotide were practically uninhibited by glucose-6-phosphate. It can thus be assumed that the equilibrium between the "active" (R) and "inactive" (T) conformations of the protein changes depending on metabolic peculiarities of a given tissue used as a source for enzyme isolation. 相似文献
6.
7.
8.
T G Sotiroudis N G Oikonomakos A E Evangelopoulos 《Biochemical and biophysical research communications》1979,86(3):674-682
Activation of phosphorylase b by AMP is stimulated by certain aliphatic and cyclic polycarboxylates. This stimulation was depended on the number and the position of the carboxyl groups, the stereochemistry and the size of the molecule, and was more pronounced at low AMP concentrations. Kinetic studies indicated that in the presence of polycarboxylates the affinity of the enzyme for AMP was enhanced, the cooperative binding of the nucleotide was removed, and the enzyme was no longer inhibited by glucose-6-phosphate. Although polycarboxylates have no effect on the sedimentation pattern of phosphorylase b in the absence of AMP, the partial association of the enzyme caused by AMP is greatly enhanced in the presence of the acids. 相似文献
9.
10.
Effects of 1,2-dimethoxyethane on the catalytic and coenzyme properties of glycogen phosphorylase 总被引:1,自引:0,他引:1
Dimethoxyethane, a good activator of phosphorylase b, has been used to study mechanisms of phosphorylase activation and the catalytic reaction. Activation can be explained best by an alteration of the allosteric equilibrium in favor of the active R conformation. Lesser effects are seen with phosphorylase a, and activation does not alter appreciably the equilibrium between the dimeric and tetrameric forms. With 20% 1,2-dimethoxyethane, the Vm value of phosphorylase b is 74% of that obtained in the presence of adenosine monophosphate. In the presence of 10% 1,2-dimethoxyethane, the Ki value for glucose inhibition is increased 3-fold, but inhibition by 1,5-gluconolactone is increased. The allosteric activation of glycogen phosphorylase results in a change in pK1 for the pH-activity profile. The formation of the dianionic form of the phosphoryl group of the coenzyme, pyridoxal phosphate, may account for this change. By analogy to the effects of anions and a change in dielectric on the acid hydroylsis of glucose 1-phosphate, it is suggested that the dianion of the coenzyme could stabilize the developing positive charge of an oxonium ion intermediate. Dimethoxyethane also affects the interaction of pyridoxal phosphate with phosphorylase. It influences the rates of both resolution and reconstitution. Good preparations of apophosphorylase a can be made by using 1,2-dimethoxyethane in the resolution medium. 相似文献
11.
12.
13.
14.
Purification and properties of yeast glycogen phosphorylase a and b 总被引:10,自引:0,他引:10
15.
Crystallization of pig skeletal phosphorylase b. Purification, physical and catalytic characterization 总被引:5,自引:0,他引:5
A new method for purification and crystallization of pig skeletal muscle phosphorylase b is presented. The ease of crystallization in the presence of 1 mM AMP and 1 mM spermine has permitted the study of some physical, chemical and enzymatic properties of the enzyme. The crystalline pig phosphorylase b gave a single band on SDS polyacrylamide gels of the same mobility as rabbit muscle phosphorylase subunit. Ultracentrifugation experiments showed that pig phosphorylase b exists in a dimeric form (S20,w = 8.4 S). No association occurred at 20 degrees C under conditions where rabbit phosphorylase b can be tetramerized; pig phosphorylase b was only 30% associated from dimer to tetramer at 13 degrees C. Pig phosphorylase b is highly stable to freezing and its specific activity did not change appreciably upon prolonged storage in the cold. Pig and rabbit phosphorylases b have comparable Vmax and Km values towards the substrate and the activator. However, there is an essential difference between the two enzymes in that pig phosphorylase b is not significantly inhibited by glucose 6-phosphate, which is a powerful inhibitor of the rabbit enzyme. Two different crystal forms of pig phosphorylase b were obtained which are small for X-ray diffraction studies. Diffusion of spermine into tetragonal crystals of rabbit phosphorylase b resulted in a difference Fourier synthesis at 3 A resolution that showed no strong indication of specific binding. 相似文献
16.
17.
Chebotareva NA Andreeva IE Makeeva VF Livanova NB Kurganov BI 《Journal of molecular recognition : JMR》2004,17(5):426-432
Self-association of phosphorylase kinase (PhK) and its interaction with glycogen (M=5500 kDa) and phosphorylase b (Phb) has been studied using analytical ultracentrifugation and turbidimetry under the conditions of molecular crowding arising from the presence of high concentrations of osmolytes. In accordance with the predictions of the molecular crowding theory, trimethylamine N-oxide (TMAO) and betaine greatly favor self-association of PhK induced by Mg2+ and Ca2+ and PhK interaction with glycogen. In contrast, proline suppresses these processes, probably, due to its specific interaction with PhK. All osmolytes tested prevented the complex formation between PhK and its physiological substrate, Phb. The specific interactions of PhK and Phb with glycogen, in the living cell, presumably is a factor allowing the negative effect of crowding on the recognition of Phb by PhK to be overcome. 相似文献
18.
Effect of glycerol on some kinetic parameters of phosphorylase b 总被引:1,自引:0,他引:1
19.
The limited proteolysis of rabbit skeletal muscle phosphorylase a and b was studied with subtilisin BPN' immoblized to Sepharose 4B. The activity of phosphorylase b is nearly resistant to subtilisin under the conditions (pH 7.0, 30 degrees C) where phosphorylase a rapidly loses its activity. The pH profile of phosphorylase a and b digestion is different. Proteolytic fragments of mol. wt 70,000 and 30,000 generated from phosphorylase a, while mol. wt 80,000 and 70,000 generated from phosphorylase b can be detected by SDS gel electrophoresis. Addition of AMP to phosphorylase b favours a conformation similar to--but not identical with--phosphorylase a as recognised by subtilisin action. 相似文献
20.
Mikleušević G Stefanić Z Narczyk M Wielgus-Kutrowska B Bzowska A Luić M 《Biochimie》2011,93(9):1610-1622
The catalytic mechanism of Escherichia coli purine nucleoside phosphorylase (PNP) is revised using site-directed mutagenesis, kinetic studies and structure determinations.The experimental evidence on the role of the particular catalytic amino acid during catalysis has not been available. Therefore, the active site mutants Arg24Ala, Asp204Ala, Asp204Asn, Arg217Ala and Asp204Ala/Arg217Ala were prepared and their kinetics and thermodynamic studies were carried out. The activity tests with natural substrates and 7-methylguanosine confirmed the earlier hypothesis, that catalysis involves protonation of the purine base at position N7 by Asp204, which is triggered by Arg217.The crystal structures of the wild type in complexes with phosphate and sulphate, respectively, and of the Arg24Ala mutant in complex with phosphate/sulphate were determined. The structural data show that previously observed conformational change is a result of the phosphate binding and its interaction with Arg24.As E. coli PNP is a promising candidate for the tumour-directed gene therapy, our results may also help to design efficient mutants useful in gene therapy. 相似文献