Generation of covalent adducts between nucleotides and proteins: applications as tools and modulators of activity

• Applicants: Professor Dr. Roger S. Goody; Professor Dr. Daniel Rauh
• Subject Area: Biological and Biomimetic Chemistry
• Term: from 2020 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 435700499

Nucleotide-binding and utilizing proteins are widespread and essential in biology. Important examples are nucleoside triphosphate-binding and -hydrolyzing proteins that do not have merely hydrolysis to nucleoside diphosphates and inorganic phosphate as their primary function, but which use the high change in free energy associated with this hydrolysis in processes such as energy conversion (e.g. motor proteins such as myosin, kinesin or dynein) or signal transduction (e.g. G-proteins or other GTPases such as those of the Ras family). To understand the functional mechanisms of such proteins, it is often necessary to characterize the in vitro and in vivo properties of their complexes with either the respective nucleoside diphosphate or nucleoside triphosphate. In order to avoid some of the possible downfalls of stabilizing a certain state by introducing point mutations that might have unintended side effects, we intend to develop methods that allow covalent binding of the respective nucleotides to the protein of interest. The nucleotide derivatives to be developed are meant, in the first instance, as tools for biochemical, biophysical and cell-biological investigations. At another level, depending on the properties of the substances to be developed, there is the additional important aim of specific inhibition of proteins involved in disease processes, in particular cancer. This goal includes targeting of the oncogenic variants G12C and G13C of Ras using cysteine-reactive warheads attached to the nucleotide that may allow locking the Ras protein in an inactive state. The work will thus include synthesis of the desired nucleotide analogues (Rauh), biophysical and biochemical characterization of their binding characteristics (Goody) and the X-ray crystallographic characterization (Rauh) to obtain molecules with the desired characteristics. One example that covalently addresses KRas G13C has already been found and is currently further optimized and characterized. At a later stage, the molecules will be tested in cellular systems in order to characterize their inhibitory potency on e.g. KRas G12C or G13C dependent cancer cell lines.

• DFG Programme: Research Grants