A 21-kDa chloroplast heat shock protein assembles into high molecular weight complexes in vivo and in Organelle.
The conservation of the carboxyl-terminal "heat shock" domain among small (sm) cytoplasmic and chloroplast heat shock proteins (HSPs) suggests that these smHSPs perform similar functions. Previous studies have established that cytoplasmic smHSPs are found in higher order structures in vivo (approximately 500 kDa). To determine if the chloroplast smHSP is found in similar complexes, we examined the size of the 21-kDa chloroplast smHSP from Pisum sativum, PsHSP21, under non-denaturing conditions. Following sedimentation of chloroplast stromal extracts on sucrose gradients PsHSP21 is detected in fractions corresponding to 10-11 S. Upon non-denaturing gel electrophoresis, PsHSP21 was detected in two high molecular mass complexes of approximately 230 and 200 kDa, in good agreement with the sucrose gradient data. These PsHSP21-containing particles were stable under different salt and Mg2+ conditions, and their integrity was not affected by 1.0% Triton X-100 or 10 mM ATP. To study assembly of the high molecular weight complexes containing PsHSP21, in vitro translated PsHSP21 was imported into chloroplasts and its size was examined. Following import into chloroplasts isolated from heat-stressed plants, greater than 50% of PsHSP21 was recovered in the higher molecular weight forms. In contrast, following import into chloroplasts isolated from control plants the protein was recovered exclusively in a 5 S (approximately 42-kDa) form. These data suggest that preexisting PsHSP21 or other heat-induced factors may be required for assembly of the higher molecular weight particles. We propose that the 10-11 S particles are the functional form of PsHSP21.
Chen
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Department of Biochemistry, University of Arizona, Tucson 85721.
Osteryoung
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K
Vierling
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D001692:Biological Transport; D015153:Blotting, Western; D002736:Chloroplasts / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007887:Fabaceae; D006360:Heat-Shock Proteins / Q000378:metabolism*; D007700:Kinetics; D008970:Molecular Weight; D010940:Plant Proteins / Q000378:metabolism*; D010946:Plants, Medicinal
D006360:Heat-Shock Proteins; D010940:Plant Proteins