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Proteomic approaches for studying alcoholism and alcohol-induced organ damage.

Hiller-Sturmhöfel S, Sobin J, Mayfield RD - Alcohol Res Health (2008)

Bottom Line: These studies have identified proteins in various brain regions whose expression is affected by alcohol.Other investigators have used proteomic approaches to identify proteins that could serve as potential biomarkers of alcohol use.Finally, interaction proteomic analyses have begun to identify proteins involved in several nerve signaling networks in the brain, which then can serve as targets for further studies on alcohol's effects.

View Article: PubMed Central - PubMed

Affiliation: CSR, Incorporated, Arlington, Virginia.

ABSTRACT
Proteomics research is concerned with the analysis of all proteins found in an organism, tissue, cell type, or cellular structure. The shotgun proteomic approach, which involves two-dimensional gel electrophoresis or liquid chromatography combined with mass spectrometry (MS), is used to identify novel proteins affected by alcohol. More targeted analyses study protein-protein interactions using such techniques as the yeast two-hybrid system, affinity chromatography, or immunoprecipitation. Finally, proteomic strategies can be combined with genomic research findings using computer analyses (i.e., in silico). All of these approaches have been used in the alcohol field. These studies have identified proteins in various brain regions whose expression is affected by alcohol. Other investigators have used proteomic approaches to identify proteins that could serve as potential biomarkers of alcohol use. Finally, interaction proteomic analyses have begun to identify proteins involved in several nerve signaling networks in the brain, which then can serve as targets for further studies on alcohol's effects. Future proteomic studies likely will shed more light on the mechanisms underlying alcohol's actions on the body.

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Related in: MedlinePlus

Schematic representation of the principles of affinity chromatography and immunoprecipitation. A) For affinity chromatography, a protein known or suspected to be involved in a protein complex (i.e., the “bait”) is attached to a solid support material to generate a stationary phase. Then, a protein extract containing potential interacting proteins is passed through this stationary phase so that complexes between the known protein and other proteins in the extract can form. By passing specific buffer solutions through the solid phase, only proteins involved in specific interactions remain and can be retrieved for further analysis. B) During immunoprecipitation, immune molecules called antibodies that interact only with a specific protein are used to pull their known target molecule out of a protein extract and with it any other proteins with which it interacts. If researchers have identified a protein that participates in a protein complex, they can produce antibodies that specifically recognize this “bait” and attach them to a solid support material. When a protein extract containing the bait and any proteins interacting with it is passed through the support, both the bait and the proteins it interacts with stick to the antibodies. Subsequently, the retained material can be retrieved and studied further.
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f3-arh-31-1-36: Schematic representation of the principles of affinity chromatography and immunoprecipitation. A) For affinity chromatography, a protein known or suspected to be involved in a protein complex (i.e., the “bait”) is attached to a solid support material to generate a stationary phase. Then, a protein extract containing potential interacting proteins is passed through this stationary phase so that complexes between the known protein and other proteins in the extract can form. By passing specific buffer solutions through the solid phase, only proteins involved in specific interactions remain and can be retrieved for further analysis. B) During immunoprecipitation, immune molecules called antibodies that interact only with a specific protein are used to pull their known target molecule out of a protein extract and with it any other proteins with which it interacts. If researchers have identified a protein that participates in a protein complex, they can produce antibodies that specifically recognize this “bait” and attach them to a solid support material. When a protein extract containing the bait and any proteins interacting with it is passed through the support, both the bait and the proteins it interacts with stick to the antibodies. Subsequently, the retained material can be retrieved and studied further.

Mentions: For affinity chromatography approaches, a protein known or suspected to be involved in a protein complex first is attached to a solid support material to generate a stationary phase that, for example, can be loaded into a glass column. Then, a protein extract containing potential interacting proteins is passed through this stationary phase, allowing the complexes between the known protein and other proteins in the extract to form (figure 3A). By passing specific buffer solutions through the columns, any proteins that do not interact specifically with the protein being studied are washed off and only the proteins involved in specific interactions remain. These then can be extracted and analyzed further in subsequent steps, such as MS analyses. The main advantage of the affinity chromatography approach over the classical yeast two-hybrid approach is that the interactions are identified in viable cells or cell extracts and, hence, do not require extensive validation (Aebersold and Mann 2003).


Proteomic approaches for studying alcoholism and alcohol-induced organ damage.

Hiller-Sturmhöfel S, Sobin J, Mayfield RD - Alcohol Res Health (2008)

Schematic representation of the principles of affinity chromatography and immunoprecipitation. A) For affinity chromatography, a protein known or suspected to be involved in a protein complex (i.e., the “bait”) is attached to a solid support material to generate a stationary phase. Then, a protein extract containing potential interacting proteins is passed through this stationary phase so that complexes between the known protein and other proteins in the extract can form. By passing specific buffer solutions through the solid phase, only proteins involved in specific interactions remain and can be retrieved for further analysis. B) During immunoprecipitation, immune molecules called antibodies that interact only with a specific protein are used to pull their known target molecule out of a protein extract and with it any other proteins with which it interacts. If researchers have identified a protein that participates in a protein complex, they can produce antibodies that specifically recognize this “bait” and attach them to a solid support material. When a protein extract containing the bait and any proteins interacting with it is passed through the support, both the bait and the proteins it interacts with stick to the antibodies. Subsequently, the retained material can be retrieved and studied further.
© Copyright Policy - public-domain
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3860448&req=5

f3-arh-31-1-36: Schematic representation of the principles of affinity chromatography and immunoprecipitation. A) For affinity chromatography, a protein known or suspected to be involved in a protein complex (i.e., the “bait”) is attached to a solid support material to generate a stationary phase. Then, a protein extract containing potential interacting proteins is passed through this stationary phase so that complexes between the known protein and other proteins in the extract can form. By passing specific buffer solutions through the solid phase, only proteins involved in specific interactions remain and can be retrieved for further analysis. B) During immunoprecipitation, immune molecules called antibodies that interact only with a specific protein are used to pull their known target molecule out of a protein extract and with it any other proteins with which it interacts. If researchers have identified a protein that participates in a protein complex, they can produce antibodies that specifically recognize this “bait” and attach them to a solid support material. When a protein extract containing the bait and any proteins interacting with it is passed through the support, both the bait and the proteins it interacts with stick to the antibodies. Subsequently, the retained material can be retrieved and studied further.
Mentions: For affinity chromatography approaches, a protein known or suspected to be involved in a protein complex first is attached to a solid support material to generate a stationary phase that, for example, can be loaded into a glass column. Then, a protein extract containing potential interacting proteins is passed through this stationary phase, allowing the complexes between the known protein and other proteins in the extract to form (figure 3A). By passing specific buffer solutions through the columns, any proteins that do not interact specifically with the protein being studied are washed off and only the proteins involved in specific interactions remain. These then can be extracted and analyzed further in subsequent steps, such as MS analyses. The main advantage of the affinity chromatography approach over the classical yeast two-hybrid approach is that the interactions are identified in viable cells or cell extracts and, hence, do not require extensive validation (Aebersold and Mann 2003).

Bottom Line: These studies have identified proteins in various brain regions whose expression is affected by alcohol.Other investigators have used proteomic approaches to identify proteins that could serve as potential biomarkers of alcohol use.Finally, interaction proteomic analyses have begun to identify proteins involved in several nerve signaling networks in the brain, which then can serve as targets for further studies on alcohol's effects.

View Article: PubMed Central - PubMed

Affiliation: CSR, Incorporated, Arlington, Virginia.

ABSTRACT
Proteomics research is concerned with the analysis of all proteins found in an organism, tissue, cell type, or cellular structure. The shotgun proteomic approach, which involves two-dimensional gel electrophoresis or liquid chromatography combined with mass spectrometry (MS), is used to identify novel proteins affected by alcohol. More targeted analyses study protein-protein interactions using such techniques as the yeast two-hybrid system, affinity chromatography, or immunoprecipitation. Finally, proteomic strategies can be combined with genomic research findings using computer analyses (i.e., in silico). All of these approaches have been used in the alcohol field. These studies have identified proteins in various brain regions whose expression is affected by alcohol. Other investigators have used proteomic approaches to identify proteins that could serve as potential biomarkers of alcohol use. Finally, interaction proteomic analyses have begun to identify proteins involved in several nerve signaling networks in the brain, which then can serve as targets for further studies on alcohol's effects. Future proteomic studies likely will shed more light on the mechanisms underlying alcohol's actions on the body.

Show MeSH
Related in: MedlinePlus