You would like to generate an antibody against the human Ras GTPase protein using a rabbit as a host. In preparation, you have prepared a sufficient quantify of the purified human protein. In two or three sentences, describe how you would generate either a monoclonal or polyclonal antibody against this protein.
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- Certain hormones, such as epinephrine, can increase the levels ofcAMP within cells. Let’s suppose you pretreat cells with or withoutepinephrine and then prepare a cell extract that contains theCREB protein.You then use an electrophoretic mobility shift assay to analyzethe ability of the CREB protein to bind to a DNA fragmentcontaining a cAMP response element (CRE). Describe what theexpected results would be.Can anyone suggest a way of increasing phosphorylation of IKK alpha (Ser176/180)? Any potent phosphatase inhibitor in combination with TNF alpha?The codon change (Gly-12 to Val-12) in human rasH that convertsit to oncogenic rasH has been associated with many types ofcancers. For this reason, researchers would like to develop drugs toinhibit oncogenic rasH. Based on your understanding of the Rasprotein, what types of drugs might you develop? In other words,what would be the structure of the drugs, and how would theyinhibit Ras protein? How would you test the efficacy of the drugs?What might be some side effects?
- An SH2-containing protein contains a mutation that changes its binding pocket such that tyrosine and phosphotyrosine bind with equal affinity. As a result, MEK activity: does not change with receptor dimerization and transautophosphorylation decreases due to changes in Raf activation increases with ligand binding-induced dimerization decreases due to allosteric inhibition of SH2-domain bindingMice and humans with inactivating mutations in the gene encoding activation-induced cytidine deaminase (AID) have an immunodeficiency disease known as ‘hyper IgM type 2’. Since AID is the enzyme that catalyzes the conversion of cytosines in the DNA to uracils, thereby initiating the process of somatic hypermutation, why do individuals with this deficiency only produce IgM antibodies?Briefly describe the following properties of the Ras GTPases: a) Size, structure and cellular localization (for structure I want to know if they are lipidated and any other unique features) , b) How are they activated and inactivated (i.e. include the GEFs and GAPs), c). Give an example of downstream effector proteins, d). Are they or could they be involved in human cancer.
- Cyclic adenosine monophosphate (CAMP) is a molecule that can act as anallosteric activator of kinase enzymes inside of the cytoplasm. Describehow CAMP molecules affect kinase enzymes?Referring to Figure 18.5, predict what would happen if the SRPRNA was unable to stimulate the GTPase activities of the GTPbindingproteins within SRP and the SRP receptor.T-cell receptors concentrate diversity in the third hypervariable region. For alpha:beta T-cell receptors, sequence diversity is heavily concentrated at the junctions formed by the rearrangement of gene segments during the generation of the expressed Va and Vb regions. The result of this organization is to position the most variable part of the T-cell receptor over a certain region of the ligand recognized by this receptor. Which region (outlined in red in figure below) indicates this part of the ligand recognized by the T-cell receptor?
- Which of the following would be expected to inhibit PKC activation by a GPCR ligand? phospholipase C inhibitor cAMP phosphodiesterase inhibitor Ras inhibitor MAP kinase inhibitorConsider the V region domains of the TCRα and TCRβ chains and the genetic loci that encode them. Which TCR chain is more genetically analogous to Ig light chain and Why?The figure from Moore (2020) shows a G protein-coupled receptor in a membrane. Note that "out" means outside the cell and "in" means facing the cytoplasm. The three amino acids "DRY" in loop I2 are required for protein targeting. What is the name of this kind of sequence, and what is its function? What protein targeting sequence is no longer included in this diagram? Why? The sequence "QXXNK" (where X is any amino acid) found in loop I3 has been identified based on its enzymatic activity. What is this activity? What particular domain would you expect to find in either Loop E1, E2, or E3? The gene that codes for this protein is a member of a family of genes that has arisen over evolutionary time. If you compared several of the genes in this family, would you expect their sequences to be most homologous (similar) in the region you describe in #3 above or #4 above? Why?