Raf Kinase Mutants
Raf Kinase Mutants - Related Videos
07:26
Generation of Organotypic Raft Cultures from Primary Human Keratinocytes
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Cited by 47 •
2012
An in vitro method to mimic in vivo epithelial differentiation is described. Many viruses target epithelial cells as part of their viral life cycle, and this method provides a means of examining virus:host interactions that more closely resembles that which occurs in vivo. This technique can be used with primary keratinocytes, established cell lines, as well as normal or diseased biopsy tissue.
10:54
Facile and Efficient Preparation of Tri-component Fluorescent Glycopolymers via RAFT-controlled Polymerization
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2015
An efficient, three-step synthesis of RAFT-based fluorescent glycopolymers, consisting of glycomonomer preparation, copolymerization, and post-modification, is demonstrated. This protocol can be used to prepare RAFT-based statistical glycopolymers with desired structures.
02:54
Protein Kinases and Phosphatases
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2020
Proteins undergo chemical modifications that trigger changes in the charge, structure, and conformation of the proteins. Phosphorylation, acetylation, glycosylation, nitrosylation, ubiquitination, lipidation, methylation, and proteolysis are various protein modifications that regulate protein activity. Such modifications are usually enzyme-driven. Protein kinases Many proteins in the cell are regulated by phosphorylation, the addition of a phosphate group. A family of enzymes called kinases...
06:09
Assaying the Kinase Activity of LRRK2 in vitro
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Cited by 3 •
2012
Leucine Rich Repeat Kinase 2 is a large multidomain kinase, mutations in which are the most common genetic cause of Parkinson's disease. Analysis of the kinase activity of this protein has proven to be a crucial tool in understanding the biology and dysfunction of this protein. In this paper, in vitro assaying of the kinase activity of LRRK2 and a selection of its mutants is described, providing an experimental system to examine phosphorylation of putative substrates and potential dysfunction...
08:05
Assaying Protein Kinase Activity with Radiolabeled ATP
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Cited by 15 •
2017
Protein kinases are highly evolved signaling enzymes and scaffolds that are critical for inter- and intracellular signal transduction. We present a protocol for measuring kinase activity through the use of radiolabeled adenosine triphosphate ([γ-32P] ATP), a reliable method to aid in elucidation of cellular signaling regulation.
FAQs
Their effects can extend beyond simply increasing kinase activity. A mutation may also change how RAF proteins interact with other pathway components or how regulatory controls constrain signaling. These alterations can produce persistent MEK–ERK pathway activity without normal growth stimuli, linking a specific molecular change to abnormal control of cell proliferation and survival.
Different substitutions can influence distinct properties of the RAF protein, including catalytic activity, protein interactions, or regulatory behavior. Consequently, two variants may produce different levels or patterns of downstream signaling. Measuring these effects helps researchers determine whether a mutation is associated with proliferation, survival, altered drug sensitivity, or resistance in a particular experimental system.
RAF function depends not only on the kinase domain but also on interactions and regulatory processes within the signaling pathway. Mutations that modify these features may keep downstream signaling active or change its response to cellular conditions. Examining these mechanisms clarifies how a variant disrupts growth control and why its biological effects may differ across models.
Researchers introduce selected variants into engineered cell systems and then examine their effects with biochemical assays and tumor models. These complementary approaches connect molecular behavior with cellular and tumor-level outcomes. Measurements can address kinase-related signaling, proliferation, survival, drug sensitivity, and resistance, helping establish how a specific mutant behaves across experimental contexts.
Engineered cells allow researchers to examine the consequences of a defined RAF alteration under controlled experimental conditions. They can test whether the variant changes proliferation or survival and assess responses to targeted inhibitors. Because the genetic change is specified, these systems help associate observed phenotypes with the mutant rather than with an unidentified background alteration.
Biochemical assays help characterize the molecular effects of a variant, while tumor models extend that analysis to tumor-related behavior. Together with engineered cell systems, they provide evidence for classifying oncogenic mutations and evaluating drug sensitivity or resistance. These results can guide development and testing of therapies intended to inhibit abnormal RAF signaling.