Human heat shock protein of apparent molecular mass 20 kDa (Hsp20) and its mutant, S16D, mimicking phosphorylation by cyclic nucleotide‐dependent protein kinases, were cloned and expressed in Escherichia coli. The proteins were obtained in a homogeneous state without utilization of urea or detergents. On size exclusion chromatography at neutral pH, Hsp20 and its S16D mutant were eluted as symmetrical peaks with an apparent molecular mass of 55–60 kDa. Chemical crosslinking resulted in the formation of dimers with an apparent molecular mass of 42 kDa. At pH 6.0, Hsp20 and its S16D mutant dissociated, and were eluted in the form of two peaks with apparent molecular mass values of 45–50 and 28–30 kDa. At pH 7.0–7.5, the chaperone activity of Hsp20 (measured by its ability to prevent the reduction‐induced aggregation of insulin or heat‐induced aggregation of yeast alcohol dehydrogenase) was similar to or higher than that of commercial α‐crystallin. Under these conditions, the S16D mutant of Hsp20 possessed lower chaperone activity than the wild‐type protein. At pH 6.0, both α‐crystallin and Hsp20 interacted with denatured alcohol dehydrogenase; however, α‐crystallin prevented, whereas Hsp20 either did not affect or promoted, the heat‐induced aggregation of alcohol dehydrogenase. The mixing of wild‐type human Hsp27 and Hsp20 resulted in a slow, temperature‐dependent formation of hetero‐oligomeric complexes, with apparent molecular mass values of 100 and 300 kDa, which contained approximately equal amounts of Hsp27 and Hsp20 subunits. Phosphorylation of Hsp27 by mitogen activated protein kinase‐activated protein kinase 2 was mimicked by replacing Ser15, 78 and 82 with Asp. A 3D mutant of Hsp27 mixed with Hsp20 rapidly formed a hetero‐oligomeric complex with an apparent molecular mass of 100 kDa, containing approximately equal quantities of two small heat shock proteins.

Oligomeric association of human small heat shock proteins HspB1, HspB5, HspB6 and HspB8 was analyzed by means of size-exclusion chromatography, analytical ultracentrifugation and chemical cross-linking. Wild-type HspB1 and Cys mutants of HspB5, HspB6 and HspB8 containing a single Cys residue in position homologous to that of Cys137 of human HspB1 were able to generate heterodimers cross-linked by disulfide bond. Cross-linked heterodimers between HspB1/HspB5, HspB1/HspB6 and HspB5/HspB6 were easily produced upon mixing, whereas formation of any heterodimers with participation of HspB8 was significantly less efficient. The size of heterooligomers formed by HspB1/HspB6 and HspB5/HspB6 was different from the size of the corresponding homooligomers. Disulfide cross-linked homodimers of small heat shock proteins were unable to participate in heterooligomer formation. Thus, monomers can be involved in subunit exchange leading to heterooligomer formation and restriction of flexibility induced by disulfide cross-linking prevents subunit exchange.

The proteins of the 14-3-3 family are universal adapters participating in multiple processes running in the cell. We describe the structure, isoform composition, and distribution of 14-3-3 proteins...

Aggregation of tau proteins followed by formation of paired helical filaments and neurofibrillary tangles is considered as a hallmark of certain neurodegenerative disorders such as different tauopathies and Alzheimer's disease (AD). Tau aggregation i