Abstract: Changes in environmental physical and chemical factors upon freeze-thawing and low temperature storage of biological samples can result in impairments of protein structures. This work specifies spontaneous and diamide-induced protein aggregations of placenta blood serum stored at -20° and -196°C during 2 years with SDS-PAGE. It was shown that storage of placenta blood serum at low temperatures did not cause any quantitative and qualitative changes in fraction distribution of proteins denatured with SDS in comparison to the native samples which were not frozen. Application of b-mercaptoethanol revealed that placenta blood serum proteins upon freeze-thawing did not form spontaneous aggregates linked by disulphide bridges. Oxidation of amino acid sulfhydryl groups induced by diamide and accompanied by high molecular aggregate formation proved to be a quite effective way for indirect estimation of structural changes in protein upon low temperature effects. In samples thawed after low temperature storage the protein aggregation with 4 mM diamide was significantly higher than in native serum. These discrepancies between native and frozen-thawed samples are stipulated by impairments of protein structure under low temperature and increased in accessibility of reactive SH-groups of proteins for oxidation with diamide. Structural changes in placenta blood serum proteins, which caused by low temperatures and revealed by elevated sensibility to diamide-induced aggregate formation, did not depend on temperature (-20° и -196°C) and storage terms (2 years and 3 weeks). They reflect protein reaction to freeze-thawing processes and could be sequence of ice crystal formation which takes place in unprotected media.
Reference: Falko O.V., Zemlianskykh N.G., Lipina O.V., Procopyuk O.S., Modification of placenta blood serum proteins under low temperature effect, Biomeditsinskaya khimiya, 2013, vol:
21. Pieper R., Gatlin C.L., Makusky A.J., Russo P.S., Schatz C.R., Miller S.S., Su Q., McGrath A.M., Estock M.A., Parmar P.P., Zhao M., Huang S.T., Zhou J., Wang F., Esquer-Blasco R., Anderson N.L., Taylor J., Steiner S. (2003) Proteomics, 3, 1345-1364. CrossRef Scholar google search