The chaperone Hsp90 is required for the appropriate regulation of numerous key signaling molecules, including the progesterone receptor (PR). Many important cochaperones bind Hsp90 through their ...tetratricopeptide repeat (TPR) domains. Two such proteins, GCUNC45 and FKBP52, assist PR chaperoning and are thought to interact sequentially with PR-Hsp90 complexes. TPR proteins bind to the C-terminal MEEVD sequence of Hsp90, but GCUNC45 has been shown also to bind to a novel site near the N-terminus. We now show that FKBP52 is also able to bind to this site, and that these two cochaperones act competitively, through Hsp90, to modulate PR activity. The N-terminal site involves noncontiguous amino acids within or near the ATP binding pocket of Hsp90. TPR interactions at this site are thus strongly regulated by nucleotide binding and Hsp90 conformation. We propose an expanded model for client chaperoning in which the coordinated use of TPR recognition sites at both N- and C-terminal ends of Hsp90 enhances its ability to coordinate interactions with multiple TPR partners.
Heat-induced Oligomerization of the Molecular Chaperone Hsp90 Chadli, Ahmed; Ladjimi, Moncef M.; Baulieu, Etienne-Emile ...
Journal of biological chemistry/The Journal of biological chemistry,
02/1999, Letnik:
274, Številka:
7
Journal Article
Recenzirano
Odprti dostop
It has been previously reported that heat shock protein 90 (Hsp90) oligomerizes at high temperatures and displays concomitantly
a novel chaperone activity (Yonehara, M., Minami, Y., Kawata, Y., ...Nagai, J., and Yahara, I. (1996) J. Biol. Chem. , 271, 2641â2645). In order to better define these oligomerization properties at high temperatures and to know whether they
are influenced by modulators of Hsp90 function, heat-induced oligomerization of highly purified dimeric Hsp90 has been investigated
over a wide range of temperature and protein concentrations by native polyacrylamide gel electrophoresis and size exclusion
chromatography. Whereas below 50â°C, the dimeric form is maintained over a large range of concentrations, at the critical
temperature of 50â°C, a sharp transition from dimeric to higher order oligomeric species takes place within minutes, in a
highly ordered process, suggesting that a conformational change, leading to the appearance of a new oligomerization site,
occurs in Hsp90 dimer. Moreover, at and above the critical temperature, the extent of oligomerization increases with Hsp90
concentration.
Formation of high order oligomers at high temperatures is sensitive to modulators of Hsp90 function. ATP and geldanamycin,
both known to bind to the same pocket of Hsp90, are inhibitors of this process, whereas molybdate, vanadate, and Nonidet P-40,
which are thought to increase surface hydrophobicity of the protein, are activators. Thus, oligomerization of Hsp90 at high
temperatures may be mediated through hydrophobic interactions that are hindered by ligands and favored by transition metal
oxyanions.
The fact that the heat-induced oligomerization of Hsp90 is affected by specific ligands that modulate its properties also
suggests that this process may be involved in cell protection during heat shock.
Mammalian endogenous carbohydrate-binding proteins (lectins) play fundamental roles in a variety of mechanisms of interactions both at the molecular and cellular levels. We have investigated the ...binding of one of them (human brain lectin) to soluble acrylamide copolymerized with derivatives of either lactose (O-beta-lactosyloxyallylallylaminoacrylamide copolymer) or D-mannose (D-alpha-mannosyloxyallylallylaminoacrylamide copolymer) in direct enzyme affinoassays, in an attempt to develop simple procedures for detection and estimation of its carbohydrate-binding activity. Biotinylated plant lectins were utilized as reference standards. Affinoassays employed the polymer dotted on nitrocellulose and the polymer coated on microtiter plates as well as detection of bound biotinylated lectin by streptavidin/horseradish peroxidase reagent. Both assays provided reproducible binding, inhibitable by specific sugars. The microtiter plate assay is well suited to sensitive detection of the negative endogenous lectin by competition with biotinylated brain lectin. We conclude that the use of derivatized acrylamide in dotting and microtiter plate assays may prove practical for detection of endogenous lectins and that such polymers may serve as model substances in the study of biological partners of these carbohydrate-binding proteins.
This article describes a method of vehicle dynamics estimation for impending rollover detection. We estimate vehicle dynamic states in presence of the road bank angle as a disturbance in the vehicle ...model using a robust observer. The estimated roll angle and roll rate are used to compute the rollover index which is based on the prediction of the lateral load transfer. In order to anticipate rollover detection, a new method is proposed to compute the time to rollover(TTR) using the load transfer ratio(LTR). The nonlinear model, deduced from the vehicle lateral and roll dynamics, is represented by a Takagi-Sugeno(T-S) fuzzy model. This representation is used to account for the nonlinearities of lateral cornering forces. The proposed T-S observer is designed with unmeasurable premise variables to cater for non-availability of the slip angles measurement. The proposed approach is evaluated using Car Sim simulator under different driving scenarios. Simulation results show good efficiency of the proposed T-S observer and the rollover detection method.
It has been previously reported that heat shock protein 90 (Hsp90) oligomerizes at high temperatures and displays concomitantly a novel chaperone activity (Yonehara, M., Minami, Y., Kawata, Y., ...Nagai, J., and Yahara, I. (1996) J. Biol. Chem., 271, 2641-2645). In order to better define these oligomerization properties at high temperatures and to know whether they are influenced by modulators of Hsp90 function, heat-induced oligomerization of highly purified dimeric Hsp90 has been investigated over a wide range of temperature and protein concentrations by native polyacrylamide gel electrophoresis and size exclusion chromatography. Whereas below 50 degreesC, the dimeric form is maintained over a large range of concentrations, at the critical temperature of 50 degreesC, a sharp transition from dimeric to higher order oligomeric species takes place within minutes, in a highly ordered process, suggesting that a conformational change, leading to the appearance of a new oligomerization site, occurs in Hsp90 dimer. Moreover, at and above the critical temperature, the extent of oligomerization increases with Hsp90 concentration. Formation of high order oligomers at high temperatures is sensitive to modulators of Hsp90 function. ATP and geldanamycin, both known to bind to the same pocket of Hsp90, are inhibitors of this process, whereas molybdate, vanadate, and Nonidet P-40, which are thought to increase surface hydrophobicity of the protein, are activators. Thus, oligomerization of Hsp90 at high temperatures may be mediated through hydrophobic interactions that are hindered by ligands and favored by transition metal oxyanions. The fact that the heat-induced oligomerization of Hsp90 is affected by specific ligands that modulate its properties also suggests that this process may be involved in cell protection during heat shock.
To understand how the molecular chaperone Hsp90 participates in conformational maturation of the estrogen receptor (ER), we analyzed the interaction of immobilized purified avian Hsp90 with mammalian ...cytosolic ER. Hsp90 was either immunoadsorbed to BF4 antibody–Sepharose or GST-Hsp90 fusion protein (GST.90) was adsorbed to glutathione-Sepharose. GST.90 was able to retain specifically ER, similarly to immunoadsorbed Hsp90. When cells were treated with estradiol and the hormone treatment was maintained during cell homogenization, binding, and washing steps, GST.90 still interacted efficiently with ER, suggesting that ER may form complexes with Hsp90 even after its activation by hormone and salt extraction from nuclei. The GST.90-ER interaction was consistently reduced in the presence of increasing concentrations of potassium chloride or when cytosolic ER-Hsp90 complexes were previously stabilized by molybdate, indicating that GST.90-ER complexes behave like cytosolic Hsp90-ER complexes. A purified thioredoxin-ER fusion protein was also able to form complexes with GST.90, suggesting that the presence of other chaperones is not required. ER was retained only by GST.90 deletion mutants bearing an intact Hsp90 N-terminal region (1–224), the interaction being more efficient when the charged region A was present in the mutant (1–334). The N-terminal fragment 1–334, devoid of the dimeric GST moiety, was also able to interact with ER, pointing to the monomeric N-terminal adenosine triphosphate binding region of Hsp90 (1–224) as the region necessary and sufficient for interaction. These results contribute to understand the Hsp90-dependent process responsible for conformational competence of ER.
This paper deals with the process of decision making in the reverse engineering mode and highlights the need for polyvalent information. Three aspects are considered. 1) Reverse engineering implies a ...preliminary assumption: having defined a desired outcome of the decision process. Defining goals on the possible outcomes is a complex, multi-actor process based on ubiquitous information. Once identified at best, several alternative scenarios may lead to the desired outcome. The first issue consists in evaluating these alternative scenarios. 2) While taking into consideration the positive consequences that the different alternatives will generate, the decision process has to allow for possible negative impacts, which are not explicitly expressed in the defined goals. We thus consider the reverse engineering process has to be bipolar and take rejections into account. 3) Finally, the simultaneous achievement (respectively, avoidance) of several goals (respectively, rejections) is not always possible and depends, in particular, on whether the actions leading to each of these goals (respectively avoiding these rejections) are compatible or not. We thus seek the "best" compatible set of actions and propose to define it as optimizing the bipolar preferences expressed on the outcomes. The approach is both graphical and logical and is focused on a case study in breadmaking technology.
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