Characterization of a sHsp of Schizosaccharomyces pombe, SpHsp15.8, and the implication of its functional mechanism by comparison with another sHsp, SpHsp16.0.
There exist two small heat shock proteins (sHsps) in the fission yeast, Schizosaccharomyces pombe (S. pombe), whose expressions are highly induced by heat stress. We have previously expressed, purified, and characterized one of the sHsps, SpHsp16.0. In this study, we examined the other sHsp, SpHsp15.8. It suppressed the thermal aggregation of citrate synthase (CS) from porcine heart and dithiothreitol-induced aggregation of insulin from bovine pancreas with very high efficiency. Almost one SpHsp15.8 subunit was sufficient to protect one protein molecule from aggregation. Like SpHsp16.0, SpHsp15.8 dissociated into small oligomers and then interacted with denatured substrate proteins. SpHsp16.0 exhibited a clear enthalpy change for denaturation occurring over 60 degrees C in differential scanning calorimetry (DSC). However, we could not observe any significant enthalpy change in the DSC of SpHsp15.8. The difference is likely to be caused by the adhesive characteristics of SpHsp15.8. The oligomer dissociation of SpHsp15.8 and SpHsp16.0 and their interactions with denatured substrate proteins were studied by fluorescence polarization analysis (FPA). Both sHsps exhibited a temperature-dependent decrease of fluorescence polarization, which correlates with the dissociation of large oligomers to small oligomers. The dissociation of the SpHsp15.8 oligomer began at about 35 degrees C and proceeded gradually. On the contrary, the SpHsp16.0 oligomer was stable up to approximately 45 degrees C, but then dissociated into small oligomers abruptly at this temperature. Interestingly, SpHsp16.0 is likely to interact with denatured CS in the dissociated state, while SpHsp15.8 is likely to interact with CS in a large complex. These results suggest that S. pombe utilizes two sHsps that function in different manners, probably to cope with a wide range of temperatures and various denatured proteins.
Department of Biotechnology and Life Science, Tokyo University of Agriculture and Technology, Koganei-shi, Tokyo 184-8588, Japan.
Chika
C
Sugino
Maya
M
Hirose
Hideki
H
Tohda
Yukiko
Y
Yoshinari
Tetsuya
T
Abe
Yuko
Y
Giga-Hama
Ryo
R
Iizuka
Masafumi
M
Shimizu
Shun-ichi
S
Kidokoro
Noriyuki
N
Ishii
Masafumi
M
Yohda
D000818:Animals; D002152:Calorimetry, Differential Scanning; D002950:Citrate (si)-Synthase / Q000378:metabolism; D005454:Fluorescence Polarization; D015966:Gene Expression Regulation, Fungal; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D050888:Heat-Shock Proteins, Small / Q000737:chemistry* / Q000378:metabolism*; D006358:Hot Temperature; D018832:Molecular Chaperones / Q000378:metabolism; D011487:Protein Conformation; D011489:Protein Denaturation; D012568:Schizosaccharomyces / Q000737:chemistry* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000378:metabolism*
D006360:Heat-Shock Proteins; D050888:Heat-Shock Proteins, Small; D018832:Molecular Chaperones; D029702:Schizosaccharomyces pombe Proteins; C491888:hsp16 protein, S pombe; D002950:Citrate (si)-Synthase