Product Pathways - Protein Folding/Stability

HSP/Chaperone Antibody Sampler Kit #9965

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Applications Key:  W=Western Blotting  IP=Immunoprecipitation  IHC-P=Immunohistochemistry (Paraffin)  IHC-F=Immunohistochemistry (Frozen)  IF-IC=Immunofluorescence (Immunocytochemistry)  F=Flow Cytometry
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  B=Bovine  Dr=Drosophila

Specificity / Sensitivity

HSP40 Antibody detects endogenous levels of total HSP40 protein. HSP60 (D307) Antibody detects endogenous levels of total HSP60 protein. HSP70 Antibody detects endogenous levels of total HSP70 protein (HSP70-Hom, HSP70-1). HSP90 (E289) Antibody detects endogenous levels of total HSP90 protein. HSF1 Antibody detects endogenous levels of total HSF1 protein. Calnexin Antibody detects endogenous levels of total calnexin protein. PDI Antibody detects endogenous levels of total PDI protein. BiP (C50B12) Rabbit mAb detects endogenous levels of total BiP protein. Each of these antibodies recognizes only its specific target.

Western Blotting

Western blot analysis of extracts from various cell types using Calnexin Antibody #2433.

Western Blotting

Western blot analysis of extracts from various cell types using PDI Antibody #2446.

Western Blotting

Western blot analysis of extracts from various cell types using BiP (C50B12) Rabbit mAb #3177.

Western Blotting

Western blot analysis of extracts from various cell types using HSF1 Antibody #4356.

Western Blotting

Western blot analysis of extracts from HeLa, C2C12 and C6 cells using HSP40 Antibody #4868.

Western Blotting

Western blot analysis of extracts from various cell types using HSP60 (D307) Antibody #4870.

Western Blotting

Western blot analysis of extracts from various cell types using HSP70 Antibody #4872.

Western Blotting

Western blot analysis of extracts from HeLa, NIH/3T3 and C6 cells using HSP90 (E289) Antibody #4875.

Source / Purification

Polyclonal antibodies are produced by immunizing rabbits with a synthetic peptide (KLH-coupled) corresponding to Glu300 of human HSP40/Hdj1, surrounding Asp307 of human HSP60, corresponding to human HSP70, surrounding Glu289 of human HSP90, corresponding to residues at the carboxy-terminus of human HSF1 protein, around Ala51 of human calnexin and around Pro329 of human PDI. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. BiP (C50B12) Rabbit mAb is produced by immunizing rabbits with a synthetic peptide (KLH-coupled) derived from the sequence around Gly584 of human BiP.

Background

HSP70 and HSP90 are molecular chaperones expressed constitutively under normal conditions to maintain protein homeostasis and are induced upon environmental stress (1). HSP70 and HSP90 interact with unfolded proteins to prevent irreversible aggregation and catalyze the refolding of their substrates in an ATP-dependent manner (1). HSP40 family proteins bind unfolded proteins and prevent their aggregation, and deliver unfolded protiens to HSP70 (2). HSP60 has primarily been known as a mitochondrial protein that is important for folding key proteins after import into the mitochondria (3). HSP60 is also present in the cytosol of many cells and is induced by stress, inflammatory and immune responses, autoantibodies correlated with Alzheimer's, coronary artery diseases, MS, and diabetes (4-7). Secretory and transmembrane proteins are synthesized on polysomes and translocate into the endoplasmic reticulum (ER) where they are often modified by the formation of disulfide bonds, amino-linked glycosylation and folding. The ER contains a pool of molecular chaperones including calnexin, BiP and protein disulfide isomerase (PDI). Calenxin is a calcium-binding protein embedded in the ER membrane that retains newly synthesized glycoproteins inside the ER to ensure proper folding and quality control (8,9). When protein folding is disturbed inside the ER, Bip synthesis is increased. Subsequently, BiP binds to misfolded proteins to prevent them from forming aggregates and assists them to refold properly (10). PDI catalyzes the formation and isomerization of disulfide bonds required to reach a proteins native state (11). Heat shock gene transcription is regulated by a familly of heat shock factors (HSFs), transcriptional activators that bind to heat shock response elements (HSEs) located upstream of all heat shock genes (12). During attenuation from the heat shock response, HSF1 is repressed by direct binding of HSP70, HSP40/Hdj-1 and HSF binding protein 1 (HSBP1) (13).

1. Nollen, E.A. and Morimoto, R.I. (2002) J. Cell Sci. 115, 2809-2816.
2. Fan, C.Y. et al. (2003) Cell Stress Chaperones 8, 309-316.
3. Jindal, S. et al. (1989) Mol Cell Biol 9, 2279-83.
4. Itoh, H. et al. (2002) Eur. J. Biochem. 269, 5931-5938.
5. Gupta, S. and Knowlton, A.A. J. Cell Mol. Med. 9, 51-58.
6. Deocaris, C.C. et al. (2006) Cell Stress Chaperones 11, 116-128.
7. Lai, H.C. et al. (2007) Am. J. Physiol. Endocrinol. Metab. 292, E292-E297.
8. Bergeron, J.J. et al. (1994) Trends Biochem. Sci. 19, 124-128.
9. Williams, D.B. (2006) J. Cell Sci. 119, 615-623.
10. Kohno, K. et al. (1993) Mol. Cell. Biol. 13, 877-890.
11. Ellgaard, L. and Ruddock, L.W. (2005) EMBO Rep. 6, 28-32.
12. Morimoto, R.I. (1998) Genes Dev. 12, 3788-3796.
13. Satyal, S.H. et al. (1998) Genes Dev. 12, 1962-1974.

Application References

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