Conformational changes and local events at the AMP site of glycogen phosphorylase b: a fluorescence temperature-jump relaxation study

Biochemistry ◽  
1978 ◽  
Vol 17 (20) ◽  
pp. 4153-4160 ◽  
Author(s):  
B. Vandenbunder ◽  
M. Dreyfus ◽  
H. Buc
Keyword(s):  
Conformational Changes ◽  
Glycogen Phosphorylase ◽  
Temperature Jump ◽  
Glycogen Phosphorylase B ◽  
Relaxation Study

scholarly journals Allosteric interactions of glycogen phosphorylase b. A crystallographic study of glucose 6-phosphate and inorganic phosphate binding to di-imidate-cross-linked phosphorylase b

1984 ◽  
Vol 218 (1) ◽  
pp. 45-60 ◽  
Author(s):  
A Lorek ◽  
K S Wilson ◽  
M S P Sansom ◽  
D I Stuart ◽  
E A Stura ◽  
...  
Keyword(s):  
Binding Site ◽  
Conformational Changes ◽  
Inorganic Phosphate ◽  
Glycogen Phosphorylase ◽  
Phosphate Binding ◽  
Binding Studies ◽  
Structural Explanation ◽  
Alpha Helices ◽  
Allosteric Effector ◽  
Glycogen Phosphorylase B

The binding to glycogen phosphorylase b of glucose 6-phosphate and inorganic phosphate (respectively allosteric inhibitor and substrate/activator of the enzyme) were studied in the crystal at 0.3 nm (3A) resolution. Glucose 6-phosphate binds in the alpha-configuration at a site that is close to the AMP allosteric effector site at the subunit-subunit interface and promotes several conformational changes. The phosphate-binding site of the enzyme for glucose 6-phosphate involves contacts to two cationic residues, Arg-309 and Lys-247. This site is also occupied in the inorganic-phosphate-binding studies and is therefore identified as a high-affinity phosphate-binding site. It is distinct from the weaker phosphate-binding site of the enzyme for AMP, which is 0.27 nm (2.7A) away. The glucose moiety of glucose 6-phosphate and the adenosine moiety of AMP do not overlap. The results provide a structural explanation for the kinetic observations that glucose 6-phosphate inhibition of AMP activation of phosphorylase b is partially competitive and highly co-operative. The results suggest that the transmission of allosteric conformational changes involves an increase in affinity at phosphate-binding sites and relative movements of alpha-helices. In order to study glucose 6-phosphate and phosphate binding it was necessary to cross-link the crystals. The use of dimethyl malondi-imidate as a new cross-linking reagent in protein crystallography is discussed.

scholarly journals Effect of Arginine on Chaperone-Like Activity of HspB6 and Monomeric 14-3-3ζ

2020 ◽  
Vol 21 (6) ◽  
pp. 2039 ◽  
Author(s):  
Valeriya V. Mikhaylova ◽  
Tatiana B. Eronina ◽  
Natalia A. Chebotareva ◽  
Vladimir V. Shubin ◽  
Daria I. Kalacheva ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Glycogen Phosphorylase ◽  
Test System ◽  
Monomeric Form ◽  
Thermal Aggregation ◽  
Scanning Calorimetry ◽  
Glycogen Phosphorylase B ◽  
Protein Chaperones ◽  
Aggregation Activity ◽  
Thermal Stability Of

The effect of protein chaperones HspB6 and the monomeric form of the protein 14-3-3ζ (14-3-3ζm) on a test system based on thermal aggregation of UV-irradiated glycogen phosphorylase b (UV-Phb) at 37 °C and a constant ionic strength (0.15 M) was studied using dynamic light scattering. A significant increase in the anti-aggregation activity of HspB6 and 14-3-3ζm was demonstrated in the presence of 0.1 M arginine (Arg). To compare the effects of these chaperones on UV-Phb aggregation, the values of initial stoichiometry of the chaperone–target protein complex (S0) were used. The analysis of the S0 values shows that in the presence of Arg fewer chaperone subunits are needed to completely prevent aggregation of the UV-Phb subunit. The changes in the structures of HspB6 and 14-3-3ζm induced by binding of Arg were evaluated by the fluorescence spectroscopy and differential scanning calorimetry. It was suggested that Arg caused conformational changes in chaperone molecules, which led to a decrease in the thermal stability of protein chaperones and their destabilization.

scholarly journals Probing the ionization state of substrate α-d-glucopyranosyl phosphate bound to glycogen phosphorylase b

1995 ◽  
Vol 308 (3) ◽  
pp. 1017-1023 ◽  
Author(s):  
I P Street ◽  
S G Withers
Keyword(s):  
Active Site ◽  
Glycogen Phosphorylase ◽  
Ph Dependence ◽  
Substrate Inhibition ◽  
19F Nmr ◽  
Ionization State ◽  
Nmr Studies ◽  
Substrate Analogues ◽  
Glycogen Phosphorylase B ◽  
Pka Value

The ionization state of the substrate alpha-D-glucopyranosyl phosphate bound at the active site of glycogen phosphorylase has been probed by a number of techniques. Values of Ki determined for a series of substrate analogue inhibitors in which the phosphate moiety bears differing charges suggest that the enzyme will bind both the monoanionic and dianionic substrates with approximately equal affinity. These results are strongly supported by 31P- and 19F-NMR studies of the bound substrate analogues alpha-D-glucopyranosyl 1-methylenephosphonate and 2-deoxy-2-fluoro-alpha-D-glucopyranosyl phosphate, which also suggest that the substrate can be bound in either ionization state. The pH-dependences of the inhibition constants K1 for these two analogues, which have substantially different phosphate pK2 values (7.3 and 5.9 respectively), are found to be essentially identical with the pH-dependence of K(m) values for the substrate, inhibition decreasing according to an apparent pKa value of 7.2. This again indicates that there is no specificity for monoanion or dianion binding and also reveals that binding is associated with the uptake of a proton. As the bound substrate is not protonated, this proton must be taken up by the proton.

scholarly journals The binding of β- and γ-cyclodextrins to glycogen phosphorylase b: Kinetic and crystallographic studies

2003 ◽  
Vol 12 (9) ◽  
pp. 1914-1924 ◽  
Author(s):  
Nikos Pinotsis ◽  
Demetres D. Leonidas ◽  
Evangelia D. Chrysina ◽  
Nikos G. Oikonomakos ◽  
Irene M. Mavridis
Keyword(s):  
Glycogen Phosphorylase ◽  
Glycogen Phosphorylase B

Effect of α-crystallin on thermal aggregation of glycogen phosphorylase b from rabbit skeletal muscle

2007 ◽  
Vol 72 (5) ◽  
pp. 518-528 ◽  
Author(s):  
A. V. Meremyanin ◽  
T. B. Eronina ◽  
N. A. Chebotareva ◽  
S. Yu. Kleimenov ◽  
I. K. Yudin ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Glycogen Phosphorylase ◽  
Rabbit Skeletal Muscle ◽  
Thermal Aggregation ◽  
Glycogen Phosphorylase B
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