Application
Western blot of APG8 fusion protein. Anti-APG8 antibody generated by immunization with recombinant yeast APG8 was tested by western blot with other anti-UBL antibodies against E.coli lysates expressing the APG8-GFP fusion protein. All UBLs possess limited homology to Ubiquitin and to each other, therefore it is important to know the degree of reactivity of each antibody against each UBL. Panel A shows total protein staining using ponceau. Panel B shows specific reaction with APG8 using a 1:4,000 and 1:8,000 dilution of GeneTex's IgG fraction of Rabbit-anti-APG8 (Yeast) followed by reaction with a 1:15,000 dilution of HRP Goat-anti-Rabbit IgGApplication
Most modifiers mature by proteolytic processing from inactive precursors (a; amino acid). Arrowheads point to the cleavage sites. Ubiquitin is expressed either as polyubiquitin or as a fusion with ribosomal proteins. Conjugation requires activating (E1) and conjugating (E2) enzymes that form thiolesters (S) with the modifiers. Modification of cullins by RUB involves SCF(SKP1/cullin-1/F-box protein) /CBC(cullin-2/elongin B/elonginC) -like E3 enzymes that are also involved in ubiquitination. In contrast to ubiquitin, the UBLs do not seem to form multi-UBL chains. UCRP(ISG15) resembles two ubiquitin moieties linked head-to-tail. Whether HUB1 functions as a modifier is currently unclear. APG12 and URM1 are distinct from the other modifiers because they are unrelated in sequence to ubiquitin. Data contributed by S.Jentsch.| Product Name | Apg8 antibody |
|---|---|
| Antibody Type | Primary Antibodies |
| Product description | Rabbit Polyclonal antibody to Apg8Ubiquitin-like proteins fall into two classes: the first class, ubiquitin-like modifiers (UBLs) function as modifiers in a manner analogous to that of ubiquitin. Examples of UBLs are SUMO, Rub1 (also called Nedd8), Apg8 and Apg12. Proteins of the second class include parkin, RAD23 and DSK2, are designated ubiquitin-domain proteins (UDPs). These proteins contain domains that are related to ubiquitin but are otherwise unrelated to each other. In contrast to UBLs, UDPs are not conjugated to other proteins. Apg8 is required for autophagy (intracellular bulk protein degradation) in yeast. Starved yeast cells take up their own cytoplasm into vacuoles through autophagic bodies. Autophagic bodies form a double-membraned structure called the autophagosome, which subsequently fuses with the vacuole/lysosome. This process similar in mammals. Two sets of genes, APG and AUT, have been identified with this process, and are responsible for two ubiquitin-like systems Apg12 and Apg8, respectively. Apg12 is synthesized in its mature form and seems to have one target, Apg5. Almost all Apg12 molecules are conjugated with Apg5. Aut2/Apg4 processes the Apg8/Aut7 system at its carboxy-terminal region. Apg8 exists in two forms, one is membrane bound through a phospholipid. Lipidation/ activation of Apg8 is mediated by Apg7 and transferred to Apg3 and finally forms a conjugate with phosphatidyl-ethanolamine (PE). Apg4 cleaves Apg8 PE, releasing Apg8 from membrane. Morphological studies show that Apg8 localizes on the membrane of intermediate structures of the autophagosome; this transient association seems to be essential for formation of the autophagosome. Both Apg12 and Apg8 are highly conserved, with apparent homologues in the worm, mammals and plants. In higher eukaryotes, Apg8 consists of a multigene family. |
| Immunogen | This purified antibody was prepared from rabbit serum after repeated immunizations with recombinant yeast APG8 protein. |
| Clonality | Polyclonal |
|---|---|
| Isotype | IgG |
| Host Species | Rabbit |
| Tested Applications | ELISAWB |
| *Optimal dilutions/concentrations should be determined by the researcher. | |
| Species Reactivity | Yeast |
| Concentration | 1mg/ml |
| Purification | Unpurified |
| Gene Synonyms | ATG8 , CVT5 , APG8 , AUT7 |
|---|---|
| Target | Apg8 |
| Cellular Localization | Membrane-associated through a lipid anchor. This association needs the 2 ubiquitin-like systems required for cytoplasm to vacuole transport and autophagy. |
Application
Western blot of APG8 fusion protein. Anti-APG8 antibody generated by immunization with recombinant yeast APG8 was tested by western blot with other anti-UBL antibodies against E.coli lysates expressing the APG8-GFP fusion protein. All UBLs possess limited homology to Ubiquitin and to each other, therefore it is important to know the degree of reactivity of each antibody against each UBL. Panel A shows total protein staining using ponceau. Panel B shows specific reaction with APG8 using a 1:4,000 and 1:8,000 dilution of GeneTex's IgG fraction of Rabbit-anti-APG8 (Yeast) followed by reaction with a 1:15,000 dilution of HRP Goat-anti-Rabbit IgG
Application
Most modifiers mature by proteolytic processing from inactive precursors (a; amino acid). Arrowheads point to the cleavage sites. Ubiquitin is expressed either as polyubiquitin or as a fusion with ribosomal proteins. Conjugation requires activating (E1) and conjugating (E2) enzymes that form thiolesters (S) with the modifiers. Modification of cullins by RUB involves SCF(SKP1/cullin-1/F-box protein) /CBC(cullin-2/elongin B/elonginC) -like E3 enzymes that are also involved in ubiquitination. In contrast to ubiquitin, the UBLs do not seem to form multi-UBL chains. UCRP(ISG15) resembles two ubiquitin moieties linked head-to-tail. Whether HUB1 functions as a modifier is currently unclear. APG12 and URM1 are distinct from the other modifiers because they are unrelated in sequence to ubiquitin. Data contributed by S.Jentsch.| Application Notes | *Optimal dilutions/concentrations should be determined by the researcher. |
|---|
| Form | Liquid |
|---|---|
| Storage Instructions | Store vial at 4° C for short-term use. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. |
| Storage Buffer | 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2 and 0.01% (w/v) Sodium Azide |
