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Mass spectrometry based structural proteomics: structural characterization of FAK/Myosin complex at molecular level

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Author(s):
Mariana Fioramonte
Total Authors: 1
Document type: Master's Dissertation
Press: Campinas, SP.
Institution: Universidade Estadual de Campinas (UNICAMP). Instituto de Química
Defense date:
Examining board members:
Fábio Cesar Gozzo; Emanuel Carrilho; Carlos Henrique Inacio Ramos
Advisor: Fábio Cesar Gozzo
Abstract

Chemical cross-linking coupled with mass spectrometry (MS) is a technique that allows the structural characterization of proteins and protein complexes, especially when these proteins are not amenable to be analyzed by high resolution techniques. The experiment is based on ther eaction of a protein or protein complex with a bifunctional reagent (cross-linking agent, ALC) followed by shotgun proteome analysis by MS. This brings to the technique all the advantages of MS, such as high sensitivity, short analysis time and ease of use. Only spatially close amino acid residues can be covalently bound by the reagent, so that the cross-linked peptides, identified by MS, serve as distance constraints between amino acids in the native structure of the protein or protein complex. The set of cross-linked peptides thus provides a set distance constraints among amino acids that can be used to build a molecular model of the protein or complex. The main objective of this work was to apply the cross-linking technique coupled to MS to map the interaction between the proteins Focal Adhesion Kinase (FAK), through its FERM domain, and myosin. This protein complex is involved in molecular signaling of the cardiac hypertrophy process. Cardiac hypertrophy is considered an adaptative mechanism of the myocardium in response to hemodynamic overload and it is associated with risk of developing heart failure, arrhythmias and sudden death. Mechamical stimuli are considered the main activator of the primary responses that lead to phenotypic changes of the myocardium in many cardiac diseases. The structural characterization of the complex FAK/myosin at the molecular level has allowed the creation of a molecular model of this complex, which was validated by mutations in the region of interaction between the proteins. This model allows the understanding the signaling processes of hypertrophy at the molecular level (AU)

FAPESP's process: 10/02628-9 - Cross-linking coupled to mass spectrometry: FERM/Miosin complex interaction mapping
Grantee:Mariana Fioramonte
Support Opportunities: Scholarships in Brazil - Master