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BIOC23H3 (23)
Lecture

BIOC23: Lec 12 - Analysis of Protein-Protein Interactions.doc

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Department
Biological Sciences
Course
BIOC23H3
Professor
Rongmin Zhao
Semester
Winter

Description
Lec 12 Analysis of ProteinProtein InteractionsMethods for proteinprotein interaction analysis1 Size exclusion chromatography2 APMSoAffinity purification coupled w mass specoTandem affinity purification TAP tag usedProtein ACalmodulin binding peptideoStepsHSP90 bound to TAPtap Protein A and Calmodulin binding peptideThe lysate is put through IgG resin where Protein A will bind the resinAfter cleavage you have HSP90 bound to only calmodulin binding peptide put this through resin of Calmodulin beadsNow youve purified the HSP90Calmodulin binding peptide protein along with HSP90s interactorsRun SDSPAGEWill get HSP90 band and several other protein bands of diff sizesCan analyze these diff proteins using mass specTrypsin digest and then tandem mass specGet sequence and then search database to determine what the proteins areoSo copurification of interactors of HSP90 and since affinity chromatography performed
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