Area C: Mechanisms and Tools

Project C01: M. Groll, E. Huber
Biochemical and crystallographic analyses of enzyme-catalysed posttranscriptional tRNA modifications

tRNA modifications are posttranscriptionally installed by specific enzymes. Close to the anticodon they affect accuracy of translation by ribosomes. N-Threonylcarbamoyladenosine is a ubiquitous modification that can undergo further cyclization as well as methylthiolation. Another sulphur-containing tRNA base is 2-thiocytidine. Since the biosynthetic pathways of both modifications are not fully understood to date, the reaction mechanisms and substrate specificities of the respective enzymes will be explored biochemically and by X-ray crystallography.

Project C02: K. Lang
Genetic code expansion tools to introduce post-translational modifications

Protein post-translational modifications (PTMs) are important for regulating diverse aspects of cellular physiology. Within this CRC we want to leverage genetic code expansion approaches - in conjunction with in vivo chemistries and enzymatic reactions – to site-specifically incorporate PTMs such as succinylation and ubiquitination into proteins in vivo. Such approaches will help us deciphering their roles – in collaboration with other groups of this CRC and in the Munich area – in nucleosome dynamics, DNA replication and transcription and protein localization. Furthermore these tools will benefit from the advantage of being generally applicable to diverse biological questions.

Project C03: H. Zipse
Acid/Base- and Redox-Properties of modified DNA Bases

Using a combination of experimental measurements and theoretical calculations several key chemical properties of modified DNA bases will be determined. This will include their acid/base profiles, their one-electron redox potentials, and bond energies for the most reactive C-H, O-H and N-H bonds. These fundamental properties will provide a basis for the rationalization of reactivity differences between canonical and modified nucleobases.

Project C04: C. Ochsenfeld
Quantum Chemical Investigations of Epigenetically Related Enzyme Mechanisms

Using newly developed, linear-scaling quantum-chemical computational methods we aim to unravel epigenetically relevant enzyme mechanisms. Up to 500-1000 atoms will be described quantum mechanically, while remaining influences on the active site by the surrounding will be described by molecular mechanics. A special focus will be on the description of the reaction mechanisms of the histone deacetylase SIRT2 and the cyclase TcdA. Further links to experimental groups within the SFB will be established based on calculating NMR shifts for a variety of biomolecular systems.

Project C05: L. Daumann
Spectroscopic and mechanistic studies of Fe/α-ketoglutarate dependent oxidases for the elucidation of demethylation mechanisms

It is the aim of this subproject to examine the key steps in the catalytic cycle of AlkBH- and TET-enzymes and to compare the local perturbations in the active site caused by the binding of different substrates with a variety of spectroscopic methods. The studies will go hand in hand with the investigation of model complexes that mimic the function of TET-enzymes. It is hoped that these studies will help to unravel the structure-reactivity relationships in TET/AlkBH-enzymes relevant in epigenetic transformations.

Project C06: F. Bracher
Synthesis and characterization of low-molecular modulators of epigenetic target enzymes

In this project novel inhibitors of histone modifying enzymes, in special type III histone deacetylases (sirtuins), will be developed. Based on our own previous work (for project A) and recently published lead structures (for project B), new, unprecedented chemotypes of inhibitors will be designed. These inhibitors will be valuable chemical tools for cooperating groups in the SFB. Optimization of the inhibitors will be fostered by contributions from other groups in the consortium.

TU München