Interaction of a small heat shock protein of the fission yeast, Schizosaccharomyces pombe, with a denatured protein at elevated temperature.
We have expressed, purified, and characterized one small heat shock protein of the fission yeast Schizosaccharomyces pombe, SpHsp16.0. SpHsp16.0 was able to protect citrate synthase from thermal aggregation at 45 degrees C with high efficiency. It existed as a hexadecameric globular oligomer near the physiological growth temperature. At elevated temperatures, the oligomer dissociated into small species, probably dimers. The dissociation was completely reversible, and the original oligomer reformed immediately after the temperature dropped. Large complexes of SpHsp16.0 and denatured citrate synthase were observed by size exclusion chromatography and electron microscopy following incubation at 45 degrees C and then cooling. However, such large complexes did not elute from the size exclusion column incubated at 45 degrees C. The denatured citrate synthase protected from aggregation was trapped by a GroEL trap mutant at 45 degrees C. These results suggest that the complex of SpHsp16.0 and denatured citrate synthase at elevated temperatures is in the transient state and has a hydrophobic nature. Analyses of the interaction between SpHsp16.0 and denatured citrate synthase by fluorescence cross-correlation spectrometry have also shown that the characteristics of SpHsp16.0-denatured citrate synthase complex at the elevated temperature are different from those of the large complex obtained after the shift to lowered temperatures.
Hirose
Maya
M
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, 2-24-16, Naka-cho, Koganei-shi, Tokyo 184-8588.
Tohda
Hideki
H
Giga-Hama
Yuko
Y
Tsushima
Reiko
R
Zako
Tamotsu
T
Iizuka
Ryo
R
Pack
Changi
C
Kinjo
Masataka
M
Ishii
Noriyuki
N
Yohda
Masafumi
M
D001671:Biochemistry / Q000379:methods; D002845:Chromatography; D002950:Citrate (si)-Synthase / Q000737:chemistry; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism; D005456:Fluorescent Dyes / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism; D006358:Hot Temperature; D008027:Light; D008854:Microscopy, Electron; D018832:Molecular Chaperones; D011485:Protein Binding; D011489:Protein Denaturation; D012542:Scattering, Radiation; D012568:Schizosaccharomyces / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013997:Time Factors
D005456:Fluorescent Dyes; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D029702:Schizosaccharomyces pombe Proteins; C491888:hsp16 protein, S pombe; D049452:Green Fluorescent Proteins; D002950:Citrate (si)-Synthase