Final Activity Report Summary - STRUCTURE OF YIDC (Structural characterisation of YidC, a novel component of the membrane proteins translocation machinery)
Biogenesis of inner membrane proteins is of central importance for all cells. In bacteria it requires targeting and insertion factors such as the signal recognition particle and the Sec translocon. YidC is an essential membrane protein, recently described as a component of the insertion machinery. Members of the YidC family (YidC/Oxa1/Alb3) exist in all three domains of life where they have multiple functions in the integration and assembly of a large variety of membrane protein complexes.
In this research project, we aimed to gain insight into the structure and function of YidC using a multidisciplinary approach. As a first essential step of the study, a rational optimisation of the cloning, production and purification protocols of E. coli YidC was established, and later successfully applied to homologous proteins. The oligomeric state and stability behaviour of the proteins were characterised in various detergent solutions. The interaction networks between YidC and the SRP and Sec translocon components were explored using complementary biophysical techniques. Broad range crystallisation screening was set up and yielded to microcrystals. The crystallisation of membrane protein represents a challenging work but many strategies were developed and planned to enhance the changes to success. The first high resolution structural data on the YidC protein family were obtained on the large periplasmic domain of E. coli YidC. The structure was determined at 1.8A resolution and revealed an unexpected fold and ligand binding site. This first breakthrough in the structure determination of the full length protein set the basis for structure function relationship studies.
In this research project, we aimed to gain insight into the structure and function of YidC using a multidisciplinary approach. As a first essential step of the study, a rational optimisation of the cloning, production and purification protocols of E. coli YidC was established, and later successfully applied to homologous proteins. The oligomeric state and stability behaviour of the proteins were characterised in various detergent solutions. The interaction networks between YidC and the SRP and Sec translocon components were explored using complementary biophysical techniques. Broad range crystallisation screening was set up and yielded to microcrystals. The crystallisation of membrane protein represents a challenging work but many strategies were developed and planned to enhance the changes to success. The first high resolution structural data on the YidC protein family were obtained on the large periplasmic domain of E. coli YidC. The structure was determined at 1.8A resolution and revealed an unexpected fold and ligand binding site. This first breakthrough in the structure determination of the full length protein set the basis for structure function relationship studies.