whereRU: Aresty Poster [#86] - Screen for Ricin A-chain mutants that have altered ribosome interaction by Aresty Posters 2009

Screen for Ricin A-chain mutants that have altered interactions with
ribosomal stalk proteins.

Contacts:
Maria Montano
mmontano@eden.rutgers.edu

Abstract:
Ricin, from the castor bean Ricinus communis, is a plant toxin and one of the most potent, lethal substances known. Ricin inhibits protein synthesis within a cell, but the molecular mechanism by which ricin kills cells and its interactions with ribosomes are not yet fully understood. The Ricin Toxin A chain (RTA) can specifically remove a conserved adenine from the ?-sarcin/ricin loop (SRL), located near the ribosomal stalk, directly affecting the elongation step of protein synthesis. If the interaction between RTA and the ribosomal stalk is critical for the depurination of the SRL and the cytotoxicity of RTA, it should be possible to isolate mutant forms of RTA that can inhibit the growth of cells solely by binding to the stalk; without depurinating the ribosome. Experimentation indicates that interaction of RTA with the ribosomal stalk contributes to its cytotoxicity in the absence of ribosome depurination. Therefore, in this study I will test the hypothesis that binding of RTA to the stalk may slow down the protein synthesis and result in a slower growth phenotype. By screening the existing RTA mutants in the laboratory I have identified one mutant that did not depurinate the ribosomes, but showed the slow-growth phenotype. I will express this RTA mutant in E. coli, purify it and study its interaction with yeast ribosomes by surface plasmon resonance techniques.