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  • Product Name
    GAPDH Antibody
  • Catalog No.
    AF0911
  • Source
    Rabbit
  • Application
    WB,ELISA
  • Reactivity
    Hm,Mk
  • UniProt
  • Mol.Wt.
    37KD
  • Concentration
    1mg/ml
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Product Information

Alternative Names:Expand▼

GAPDH, A1 40 kd subunit, Activator 1 40 kd subunit, G3PD, GAPD, G3pdh, Rfc40, Rf-c 40 kd subunit

Applications:

WB: 1:5000~1:50000, ELISA(peptide) 1:20000-1:40000

Reactivity:

Human,Monkey

Source:

Rabbit

Clonality:

Polyclonal

Purification:

The antiserum was purified by peptide affinity chromatography using SulfoLink™ Coupling Resin (Thermo Fisher Scientific).

Specificity:

GAPDH antibody detects endogenous levels of total GAPDH protein from human and monkey.Is unsuitable for MOUSE and RAT,not yet tested in other species.

Format:

Liquid

Concentration:

1mg/ml

Storage Condition and Buffer:

Rabbit IgG in phosphate buffered saline, pH 7.4, 150mM NaCl, 0.02% sodium azide and 50% glycerol.

Immunogen Information

Immunogen:

A synthesized peptide derived from human GAPDH.

Uniprot:



>>Visit The Human Protein Atlas

Gene id:

Molecular Weight:

Observed Mol.Wt.: 37KD.
Predicted Mol.Wt.: 37kDa.

Subcellular Location:

Cytoplasm > cytosol. Nucleus. Cytoplasm > perinuclear region. Membrane. Translocates to the nucleus following S-nitrosylation and interaction with SIAH1, which contains a nuclear localization signal (By similarity). Postnuclear and Perinuclear regions.

Description:

Glyceraldehyde 3 phosphate dehydrogenase (GAPDH) is well known as one of the key enzymes involved in glycolysis. GAPDH is constitutively abundant expressed in almost cell types at high levels, therefore antibodies against GAPDH are useful as loading controls for Western Blotting. Some pathology factors, such as hypoxia and diabetes, increased or decreased GAPDH expression in certain cell types.

Sequence:
        10         20         30         40         50
MGKVKVGVNG FGRIGRLVTR AAFNSGKVDI VAINDPFIDL NYMVYMFQYD
60 70 80 90 100
STHGKFHGTV KAENGKLVIN GNPITIFQER DPSKIKWGDA GAEYVVESTG
110 120 130 140 150
VFTTMEKAGA HLQGGAKRVI ISAPSADAPM FVMGVNHEKY DNSLKIISNA
160 170 180 190 200
SCTTNCLAPL AKVIHDNFGI VEGLMTTVHA ITATQKTVDG PSGKLWRDGR
210 220 230 240 250
GALQNIIPAS TGAAKAVGKV IPELNGKLTG MAFRVPTANV SVVDLTCRLE
260 270 280 290 300
KPAKYDDIKK VVKQASEGPL KGILGYTEHQ VVSSDFNSDT HSSTFDAGAG
310 320 330
IALNDHFVKL ISWYDNEFGY SNRVVDLMAH MASKE

Background

Function:

Has both glyceraldehyde-3-phosphate dehydrogenase and nitrosylase activities, thereby playing a role in glycolysis and nuclear functions, respectively. Participates in nuclear events including transcription, RNA transport, DNA replication and apoptosis. Nuclear functions are probably due to the nitrosylase activity that mediates cysteine S-nitrosylation of nuclear target proteins such as SIRT1, HDAC2 and PRKDC. Modulates the organization and assembly of the cytoskeleton. Facilitates the CHP1-dependent microtubule and membrane associations through its ability to stimulate the binding of CHP1 to microtubules (By similarity). Glyceraldehyde-3-phosphate dehydrogenase is a key enzyme in glycolysis that catalyzes the first step of the pathway by converting D-glyceraldehyde 3-phosphate (G3P) into 3-phospho-D-glyceroyl phosphate. Component of the GAIT (gamma interferon-activated inhibitor of translation) complex which mediates interferon-gamma-induced transcript-selective translation inhibition in inflammation processes. Upon interferon-gamma treatment assembles into the GAIT complex which binds to stem loop-containing GAIT elements in the 3'-UTR of diverse inflammatory mRNAs (such as ceruplasmin) and suppresses their translation.

Post-translational Modifications:

S-nitrosylation of Cys-152 leads to interaction with SIAH1, followed by translocation to the nucleus (By similarity). S-nitrosylation of Cys-247 is induced by interferon-gamma and LDL(ox) implicating the iNOS-S100A8/9 transnitrosylase complex and seems to prevent interaction with phosphorylated RPL13A and to interfere with GAIT complex activity.ISGylated.Sulfhydration at Cys-152 increases catalytic activity.Oxidative stress can promote the formation of high molecular weight disulfide-linked GAPDH aggregates, through a process called nucleocytoplasmic coagulation. Such aggregates can be observed in vivo in the affected tissues of patients with Alzheimer disease or alcoholic liver cirrhosis, or in cell cultures during necrosis. Oxidation at Met-46 may play a pivotal role in the formation of these insoluble structures. This modification has been detected in vitro following treatment with free radical donor (+/-)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide. It has been proposed to destabilize nearby residues, increasing the likelihood of secondary oxidative damages, including oxidation of Tyr-45 and Met-105. This cascade of oxidations may augment GAPDH misfolding, leading to intermolecular disulfide cross-linking and aggregation.

Subcellular Location:

Nucleus;Cytosol;Cytoskeleton;

Extracellular region or secreted Cytosol Plasma membrane Cytoskeleton Lysosome Endosome Peroxisome ER Golgi apparatus Nucleus Mitochondrion Manual annotation Automatic computational assertionGraphics by Christian Stolte

Subunit Structure:

Homotetramer. Interacts with TPPP; the interaction is direct. Interacts (when S-nitrosylated) with SIAH1; leading to nuclear translocation. Interacts with RILPL1/GOSPEL, leading to prevent the interaction between GAPDH and SIAH1 and prevent nuclear translocation. Interacts with CHP1; the interaction increases the binding of CHP1 with microtubules. Associates with microtubules (By similarity). Interacts with EIF1AD, USP25, PRKCI and WARS. Interacts with phosphorylated RPL13A; inhibited by oxidatively-modified low-densitity lipoprotein (LDL(ox)). Component of the GAIT complex. Interacts with FKBP6; leading to inhibit GAPDH catalytic activity.

Similarity:

The [IL]-x-C-x-x-[DE] motif is a proposed target motif for cysteine S-nitrosylation mediated by the iNOS-S100A8/A9 transnitrosylase complex.Belongs to the glyceraldehyde-3-phosphate dehydrogenase family.

Research Fields

Research Fields:

· Environmental Information Processing > Signal transduction > HIF-1 signaling pathway.(View pathway)
· Human Diseases > Neurodegenerative diseases > Alzheimer's disease.
· Metabolism > Global and overview maps > Biosynthesis of amino acids.
· Metabolism > Global and overview maps > Carbon metabolism.
· Metabolism > Carbohydrate metabolism > Glycolysis / Gluconeogenesis.
· Metabolism > Global and overview maps > Metabolic pathways.

Emodin targets mitochondrial cyclophilin D to induce apoptosis inHepG2 cellsLing Zhang, Dian He, Kun Li, Hongli Liu, Baitao Wang, Lifang Zheng*, Jiazhong Li
Western blot analysis of GAPDH Antibody expression in JK, Hela, 293, A549, cos7 and MCF-7 cells lysates.
Western blot analysis of Hela (1), Jurket (2), and COS-7(3) with GAPDH antibody(1;50000),The lane on the left is treated with the antigen-specific peptide.

Reference Citations:

1). Zhang L et al. Emodin targets mitochondrial cyclophilin D to induce apoptosis in HepG2 cells. Biomed Pharmacother 2017 Jun;90:222-228 (PubMed: 28363167)

Application: WB    Species: human;    Sample:Not available;

Fig. 3. Effects of ROS and ERK on CypD expression. Cells were treated with emodin in the absence or presence of 5mM CsA or 10 ng/mL EGF for 48 h, or 5 mM NAC for 6 h. Protein expression was determined by western blots. (A) CypD expression in the absence or presence of CsA. (B) p-ERK expression induced by emodin. (C) CypD expression in the absence or presence of EGF. (D) CypD expression in the absence or presence of NAC. (E) Determination of emodin-induced cytotoxicity activity in the absence or presence of EGF or NAC by MTT assay. Values are expressed as the means  SD; n = 3, * p < 0.05 versus control.


2). Hui-Fang Wang et al. Crocetin inhibits the proliferation, migration and TGF-β 2-induced epithelial-mesenchymal transition of retinal pigment epithelial cells . EUR J PHARMACOL 2017 Nov;815:391-398

3). Wang HF et al. Crocetin inhibits the proliferation, migration and TGF-β2-induced epithelial-mesenchymal transition of retinal pigment epithelial cells. Eur J Pharmacol 2017 Nov 15;815:391-398 (PubMed: 28970011)

Application: WB 1/5000    Species: human;    Sample: ARPE-19;

Figure 4. Crocetin inhibits TGF-β2-induced EMT. After 1 h of pretreatment with crocetin, ARPE-19 cells used for EMT assay were stimulated with or without recombinant human TGF-β2 for up to 24 or 48 h. (A) Phase contrast photomicrographs of confluent cultures of cells were captured after treatment for 48 h. Scale bar: 200 μm. (B) Western blot analysis levels of of ZO-1, E-cadherin, Vimentin, α-SMA and the housekeeping protein GAPDH in the lysates of ARPE-19 cells after treatment for 48 h. *P< 0.05, **P< 0.01, ***P< 0.001. The data are presented as the mean ± S.D. (n = 3/group).


4). Zhao H et al. CCL17-CCR4 axis promotes metastasis via ERK/MMP13 pathway in bladder cancer. J Cell Biochem 2018 Sep 19 (PubMed: 30230587)

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Catalog Number :

AF0911-BP

Price/Size :

$200/1mg.
Tips: For phospho antibody, we provide phospho peptide(0.5mg) and non-phospho peptide(0.5mg).

Function :

Blocking peptides are peptides that bind specifically to the target antibody and block antibody binding. These peptide usually contains the epitope recognized by the antibody. Antibodies bound to the blocking peptide no longer bind to the epitope on the target protein. This mechanism is useful when non-specific binding is an issue, for example, in Western blotting (immunoblot) and immunohistochemistry (IHC). By comparing the staining from the blocked antibody versus the antibody alone, one can see which staining is specific; Specific binding will be absent from the western blot or immunostaining performed with the neutralized antibody.

Format and storage :

Synthetic peptide was lyophilized with 100% acetonitrile and is supplied as a powder. Reconstitute with 0.1 ml DI water for a final concentration of 1 mg/ml.The purity is >90%,tested by HPLC and MS.Storage Maintain refrigerated at 2-8°C for up to 6 months. For long term storage store at -20°C.

Precautions :

This product is for research use only. Not for use in diagnostic or therapeutic procedures.

IMPORTANT: For western blots, incubate membrane with diluted antibody in 5% w/v milk , 1X TBS, 0.1% Tween®20 at 4°C with gentle shaking, overnight.