Graduate Students Mini Symposium X - 2024

13:15 h Chris Wilson - AG Bode
“New tools for engineering combinatorial peptide libraries from hybrid non-ribosomal peptide synthetases”

Nonribosomal peptide synthetases (NRPS) are a valuable source of bioactive compounds, making them a prime target for engineering new compounds with enhanced bioactivity. Several methods, including the XU, XUC, and XUT approaches, have been developed for NRPS engineering. Here, we present a novel method that combines these techniques. To assess this approach, we tested multiple fusion sites to reconstitute the xenotetrapeptide synthetase (XtpS). Additionally, we established a functional tripartite split system using our model XtpS system. Finally, we successfully produced a combinatorial peptide library from a variety of hybrid NRPS.

13:45 h Nurseda Yilmaz - MPRG Höfer
"RNAylated proteins as a platform to develop next-generation RNA therapeutics"

RNA therapeutics hold great promise but face challenges like instability in eukaryotic systems, where exogenous RNA is degraded. RNAylation, a novel RNA modification, covalently attaches RNA to proteins enzymatically, enhancing RNA stability by protecting it from nuclease degradation. We developed a modular platform using RNAylation to generate RNA-protein conjugates with exchangeable nucleic acid and protein components tailored for specific applications. This platform enables precise control over the localization and function of RNA-protein conjugates, with an efficient purification strategy for therapeutic use. We also demonstrated successful delivery of RNAylated proteins to eukaryotic cells, establishing a foundation for RNA-based therapeutics.

14:15 h Fidel Ramirez - AG Schuller
"The machinery shaping methanogenesis and F₄₃₀"

Methyl-coenzyme M reductase (MCR) serves as the enzyme responsible for the production of nearly all biologically generated methane. Deeply within its active site, MCR comprises coenzyme F430, a porphyrin-based cofactor with a central nickel ion that is active exclusively in the Ni(I) state. How methanogens achieve the redox potential of the Ni(I)/Ni(II) couple (E°’ = -650 mV) for the reductive activation of F430 remains as a major question to understand such ancient bioenergetic system. In the mcrBDCGA operon of M. maripaludis, mcrC encodes for a chaperone protein involved in the activation of MCR. By integrating a Twin-Strep (TS) tag in the N-terminus of McrC, it was possible to co-purify MCR along with the previously uncharacterized methanogenic marker proteins Mmp7, Mmp17, Mmp3 and the A2 component. This complex can activate MCR in vitro in an ATP-dependent manner, enabling the production of methane. By carrying out redox-controlled cryoEM, a 2.1 Å structure of the MCR activation complex was obtained. Strikingly, a series of metallocofactors embedded in the activation complex shape an electron transfer pathway towards F430. Overall, our structural and biochemical evidence offer novel insights on the ATP-dependent activation mechanism of MCR.