Probing the mechanism of a molecular machine by engineering a light-switchable group II chaperonin

• Applicant: Dr. Daniel Hörsch
• Subject Area: Biochemistry
• Term: from 2011 to 2013
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 197676722

Groups II chaperonins are molecular machines employing the energy of ATP hydrolysis to mediate protein folding in eukaryotes and archea via a still largely enigmatic mechanism. I propose an engineering approach towards elucidating chaperonin mechanism with two interrelated goals: First, to test functional hypotheses derived from recent high-resolution structures of group II chaperonins in multiple conformations; and second, to convert a model chaperonin, the biochemically tractable archeal Mm-cpn, from an ATP- to a light-driven machine. Methodologically, I will combine computer-aided design borrowing techniques from robotics to model conformational changes of protein machines (developed in the host laboratory), chaperonin biochemistry and structural analysis pioneered by our collaborators (Frydman, Chiu and Adams) and chemical biology to engineer light-controlled protein function. Azobenzene compounds change their end-to-end distance reversibly when exposed to light of different wavelength. I will attach thiol-reactive azobenzene cross-linkers to engineered pairs of cysteines on Mm-cpn at predicted critical sites to photo-switch its conformational transitions. If successful, controlling key steps in the chaperonin conformational cycle using the temporal precision of light should enable insights into chaperonin function at a resolution previously not possible. Both dissecting mechanism using such means and engineering a light-driven protein machine would be a first.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Professorin Dr. Tanja Kortemme