UV-irradiated BSA: The details of aggregation kinetics and structural rearrangements.

UV-irradiation is a stress factor for proteins, leading to disruption of their native structure. Test systems based on UV-irradiated proteins are relevant for researchers, as they allow working directly with damaged protein molecules, which can be important when studying the properties and mechanisms of action of various antiaggregation agents. The study of UV-irradiated proteins can also have applied significance, including medical.

Effects of osmolytes under crowding conditions on the properties of muscle glycogen phosphorylase b.

The study of the relationship between the activity and stability of enzymes under crowding conditions in the presence of osmolytes is important for understanding the functioning of a living cell. The effect of osmolytes (trehalose and betaine) on the secondary and tertiary structure and activity of muscle glycogen phosphorylase b (Phb) under crowding conditions created by PEG 2000 and PEG 20000 was investigated using dynamic light scattering, differential scanning calorimetry, circular dichroism spectroscopy, fluorimetry and enzymatic activity assay. At 25 °C PEGs increased Phb activity, but PEG 20000 to a greater extent.

Change in the Kinetic Regime of Aggregation of Yeast Alcohol Dehydrogenase in the Presence of 2-Hydroxypropyl-β-cyclodextrin.

Chemical chaperones are low-molecular-weight compounds that suppress protein aggregation. They can influence different stages of the aggregation process-the stage of protein denaturation, the nucleation stage and the stage of aggregate growth-and this may lead to a change in the aggregation kinetic regime. Here, the possibility of changing the kinetic regime in the presence of a chemical chaperone 2-hydroxypropyl-β-cyclodextrin (2-HP-β-CD) was investigated for a test system based on the thermally induced aggregation of yeast alcohol dehydrogenase (yADH) at 56 °C.

Effect of Chemical Chaperones on the Stability of Proteins during Heat- or Freeze-Thaw Stress.

The importance of studying the structural stability of proteins is determined by the structure-function relationship. Protein stability is influenced by many factors among which are freeze-thaw and thermal stresses. The effect of trehalose, betaine, sorbitol and 2-hydroxypropyl-β-cyclodextrin (HPCD) on the stability and aggregation of bovine liver glutamate dehydrogenase (GDH) upon heating at 50 °C or freeze-thawing was studied by dynamic light scattering, differential scanning calorimetry, analytical ultracentrifugation and circular dichroism spectroscopy.

The Effect of Chemical Chaperones on Proteins with Different Aggregation Kinetics.

Formation and accumulation of protein aggregates adversely affect intracellular processes in living cells and are negative factors in the production and storage of protein preparations. Chemical chaperones can prevent protein aggregation, but this effect is not universal and depends on the target protein structure and kinetics of its aggregation. We studied the effect of betaine (Bet) and lysine (Lys) on thermal aggregation of muscle glycogen phosphorylase b (Phb) at 48°C (aggregation order, n = 0.

Effect of Trehalose on Oligomeric State and Anti-Aggregation Activity of αB-Crystallin.

αB-Crystallin (αB-Cr), one of the main crystalline lens proteins, along with other crystallins maintains lens transparency suppressing protein aggregation and thus preventing cataractogenesis. αB-Cr belongs to the class of molecular chaperones; being expressed in many tissues it has a dynamic quaternary structure, which is essential for its chaperone-like activity. Shift in the equilibrium between ensembles of oligomers of different size allows regulating the chaperone activity.

Effect of Betaine and Arginine on Interaction of αB-Crystallin with Glycogen Phosphorylase .

Protein-protein interactions (PPIs) play an important role in many biological processes in a living cell. Among them chaperone-client interactions are the most important. In this work PPIs of αB-crystallin and glycogen phosphorylase (Ph) in the presence of betaine (Bet) and arginine (Arg) at 48 °C and ionic strength of 0.

Combined action of chemical chaperones on stability, aggregation and oligomeric state of muscle glycogen phosphorylase b.

Chemical chaperones are a class of small molecules, which enhance protein stability, folding, inhibit protein aggregation, and are used for long-term storage of therapeutic proteins. The combined action of chemical chaperones trehalose, betaine and lysine on stability, aggregation and oligomeric state of muscle glycogen phosphorylase b (Phb) has been studied. Dynamic light scattering data indicate that the affinity of trehalose to Phb increased in the presence of betaine or lysine at both stages (stage of nucleation and aggregate growth) of enzyme aggregation at 48 °C, in contrast, the affinity of betaine to the enzyme in the presence of lysine remained practically unchanged.

Effect of arginine on stability and aggregation of muscle glycogen phosphorylase b.

Arginine (Arg) is frequently used in biotechnology and pharmaceutics to stabilize protein preparations. When using charged ions like Arg, it is necessary to take into account their contribution to the increase in ionic strength, in addition to the effect of Arg on particular processes occurring under the conditions of constancy of ionic strength. Here, we examined contribution of ionic strength (0.

Chaperone-Like Activity of HSPB5: The Effects of Quaternary Structure Dynamics and Crowding.

Small heat-shock proteins (sHSPs) are ATP-independent molecular chaperones that interact with partially unfolded proteins, preventing their aberrant aggregation, thereby exhibiting a chaperone-like activity. Dynamics of the quaternary structure plays an important role in the chaperone-like activity of sHSPs. However, relationship between the dynamic structure of sHSPs and their chaperone-like activity remains insufficiently characterized.

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

The effect of protein chaperones HspB6 and the monomeric form of the protein 14-3-3ζ (14-3-3ζ) on a test system based on thermal aggregation of UV-irradiated glycogen phosphorylase (UV-Ph) 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 14-3-3ζ was demonstrated in the presence of 0.

Comparative effects of trehalose and 2-hydroxypropyl-β-cyclodextrin on aggregation of UV-irradiated muscle glycogen phosphorylase b.

Chemical chaperones are a class of small molecules which enhance folding and prevent aggregation of proteins. Investigation of their effects on the processes of protein aggregation is of importance for further understanding of implication of protein aggregation in neurodegenerative diseases, as well as for solving biotechnological tasks. The effects of chemical chaperones trehalose and 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) on the kinetics of aggregation of UV-irradiated muscle glycogen phosphorylase b (UV-Phb) at 37 °C have been studied.

Kinetic regime of Ca and Mg-induced aggregation of phosphorylase kinase at 40 °C.

Many functions of phosphorylase kinase (PhK) are regulated by Ca and Mg ions. Ca and Mg ions stimulate activity of PhK, induce the changes in the tertiary and quaternary structure of the hexadecameric enzyme molecule, provoke association/aggregation of PhK molecules, enhance PhK binding to glycogen. To establish the kinetic regime of Ca and Mg-induced aggregation of PhK from rabbit skeletal muscles at 40 °C, in the present work the kinetics of aggregation was studied at various protein concentrations using the dynamic light scattering.

Oligomeric state of αB-crystallin under crowded conditions.

Small heat shock proteins (sHsps) are molecular chaperones preventing protein aggregation. Dynamics of quaternary structure plays an important role in the chaperone-like activity of sHsps. However, an interrelation between the oligomeric state and chaperone-like activity of sHsps remains insufficiently characterized.

Effect of ionic strength and arginine on aggregation of UV-irradiated muscle glycogen phosphorylase b.

In this work the effect of ionic strength and arginine on the kinetics of aggregation of UV-irradiated muscle glycogen phosphorylase b (UV-Phb) was studied using dynamic light scattering at 37 °C at various ionic strengths (0.02-0.7 M).

A thermal after-effect of UV irradiation of muscle glycogen phosphorylase b.

Different test systems are used to characterize the anti-aggregation efficiency of molecular chaperone proteins and of low-molecular-weight chemical chaperones. Test systems based on aggregation of UV-irradiated protein are of special interest because they allow studying the protective action of different agents at physiological temperatures. The kinetics of UV-irradiated glycogen phosphorylase b (UV-Phb) from rabbit skeletal muscle was studied at 37°C using dynamic light scattering in a wide range of protein concentrations.

Mechanism of aggregation of UV-irradiated glycogen phosphorylase b at a low temperature in the presence of crowders and trimethylamine N-oxide.

To characterize the initial stages of protein aggregation, the kinetics of aggregation of UV-irradiated glycogen phosphorylase b (UV-Phb) was studied under conditions when the aggregation proceeded at a low rate (10°C, 0.03M Hepes buffer, pH6.8, containing 0.

Kinetic regime of thermal aggregation of holo- and apoglycogen phosphorylases b.

To characterize the role of pyridoxal 5'-phosphate in stabilization of the conformation of muscle glycogen phosphorylase b (Phb), the mechanism of thermal aggregation for holo- and apoforms of Phb has been studied using dynamic light scattering. The order of aggregation with respect to the protein (n) for aggregation of holoPhb at 48°C is equal to 0.5 suggesting that the dissociative mechanism of denaturation is operative and denaturation is followed by rapid aggregation stage.

Checking for reversibility of aggregation of UV-irradiated glycogen phosphorylase b under crowding conditions.

It is believed that the initial stages of protein aggregation are reversible and can be reversed by simple dilution, whereas prolonged exposure to factors responsible for denaturing proteins (for example, to elevated temperatures) results in the formation of irreversible aggregates. A new approach has been developed to discriminate the stage of the formation of reversible aggregates. Aggregation of UV-irradiated glycogen phosphorylase b (UV-Phb) was studied at 10, 25 and 37 °C in the presence of crowders (polyethylene glycol and Ficoll-70) using dynamic light scattering and analytical ultracentrifugation (pH 6.

Dual effect of arginine on aggregation of phosphorylase kinase.

Arginine is widely used in biotechnology as a folding enhancer and aggregation suppressor. However, its action on the stability of complexly organized oligomeric proteins, on the one hand, and its role in the formation of supramolecular structures, on the other hand, are poorly known. The investigation is concerned with the effects of arginine on protein-protein interactions using phosphorylase kinase (PhK) as an example.