Heat induced protein denaturation in the particulate fraction of hela S3 cells: Effect of thermotolerance

Abstract

In this study we investigated the effect of heat on the proteins of the particulate fraction (PF) of HeLa S3 cells using electron spin resonance (ESR) and thermal gel analysis (TGA). ESR detects overall conformational changes in proteins, while TGA detects denaturation (aggregation due to formation of disulfide bonds) in specific proteins. For ESR measurements the -SH groups of the proteins were labelled with a maleimido bound spin label (4-maleimido-tempo). The sample was heated inside the ESR spectrometer at a rate of 1°/min. ESR spectra were made every 2–3° between 20° and 70°. In the PF of untreated cells conformational changes in proteins were observed in three temperature streches: between 38 and 44° (transition A, T_A); between 47 and 53° (transition B, T_B); and above 58° (transition C, T_C). With TGA, using the same heating rate, we identified three proteins (55, 70, and 90kD) which denatured during T_B. No protein denaturation was observed during T_A, while during T_C denaturation of all remaining proteins in the PF occurred. When the ESR and TGA measurements were done with the PF of (heat-induced) thermotolerant ceils, T_A was unchanged while T_B and T_C started at lated with the degree of thermotolerance that was induce in the cells. These results suggest that protection against heat-induced denaturation of proteins in the PF in involved in heat induced thermotolerance. © 1992 Wiley-Liss, Inc