Thermal denaturation and aggregation of apoform of glycogen phosphorylase b. Effect of crowding agents and chaperones

Abstract

The effect of protein and chemical chaperones and crowders on thermal stability and aggregation of apoform of rabbit muscle glycogen phosphorylase b (apoPhb) has been studied at 37°C. Proline suppressed heat-induced loss in ability of apoPhb to reconstitution at 37°C, whereas α-crystallin did not reveal a protective action. To compare the antiaggregation activity of intact and crosslinked α-crystallins, an adsorption capacity (AC) of a protein chaperone with respect to a target protein was estimated. This parameter is a measure of the antiaggregation activity. Crosslinking of α-crystallin results in 11-fold decrease in the initial AC. The nonlinear character of the relative initial rate of apoPhb aggregation versus the [intact α-crystallin]/[apoPhb] ratio plot is indicative of the decrease in the AC of α-crystallin with increasing the [α-crystallin]/[apoPhb] ratio and can be interpreted as an evidence for dynamic chaperone structure and polydispersity of α-crystallin-target protein complexes. As for chemical chaperones, a semisaturation concentration of the latter was used as a characteristic of the antiaggregation activity. A decrease in the semisaturation concentration for proline was observed in the presence of the crowders (polyethylene glycol and Ficoll-70).

Effect of crowding on several stages of protein aggregation in test systems in the presence of α-crystallin

“…The reason of the appearance of the sensitivity of the nucleation stage and the stages of clusterization of nuclei to crowding is incorporation of ␣-crystallin particles in nuclei and their clusters. ␣-Crystallin particles possess dynamic structure and show a capacity for intermolecular exchange of subunits and partial dissociation into suboligomeric forms when interacting with target protein [9,15,[48][49][50][51][52][53][54][55][56]. Carver and coworkers showed that the rate of subunit exchange of ␣A-crystallin decreased under crowding conditions [57].…”

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

“…The anti-aggregation activity of protein chaperones can be characterized by the value of the initial stoichiometry of the chaperone-target protein complex (S 0 ) and can be determined from the initial linear part of the dependence of v 0 on the [chaperone]/[UV-Phb] ratio [41]:…”

Polyols (Glycerol and Ethylene glycol) mediated amorphous aggregate inhibition and secondary structure restoration of metalloproteinase-conalbumin (ovotransferrin)

“…(apoPhb) of rabbit origin [12], while Aquaporin 0 (AQP0) aggregates at 60 • C [13], bovine hemoglobin aggregates at ∼70 • C [14], recombinant human interferon alpha2b (rhIFN␣2b) aggregates at 50 • C [15] while SARS-associated corona virus (SARS-CoV) protein aggregates at most extreme condition i.e. boiling temperature [16].…”