Biology subjects

Bruenle, S.

Publications and source records attributed to Bruenle, S..

3 recordsLinked to original sources

bioRxiv2025

Visualizing the mechanism of quinol oxidation and inhibition of a bd-type oxidase using cryo-EM

Cytochrome bd is a prokaryotic terminal oxidase recognized as an antibiotic target against various pathogens. Despite its critical role in respiration, failure to capture the mechanism of enzyme catalysis and inhibition prohibits structure guided drug discovery. Here, we present cryo-electron microscopy structures of Escherichia coli cytochrome bd-I in monomeric and dimeric forms, along all stages of quinol turn-over and in an inhibitor-bound state. We identify a dynamic Q-loop lid that undergoes a disorder-to-order transition upon substrate binding to the dimer, completing the active site and enabling catalysis. Structure-guided mutagenesis confirms Tyr243 and Arg298 as essential catalytic residues unique to long Q-loop oxidases, highlighting evolutionary divergence from short Q-loop variants. Inhibition by Aurachin D triggers refolding of the active site, occluding substrate access via a conserved Asp239-mediated mechanism. The structural and mechanistic insights presented here establish a comprehensive framework, opening new ways for drug discovery.

biochemistry↗

bioRxiv2024

Structural insights into the opening mechanism of C1C2 channelrhodopsin

Channelrhodopsins, light-gated cation channels, enable precise control of neural cell depolarization or hyperpolarization with light in the field of optogenetics. This study integrates time-resolved serial crystallography and atomistic molecular dynamics (MD) simulations to resolve the structural changes during C1C2 channelrhodopsin activation. Our observations reveal that within the crystal environment, C1C2 predominantly remains in a light-activated state with characteristics of the M390 intermediate. Here, rearrangement of retinal within its binding pocket partially opens the central gate towards the extracellular vestibule. These structural changes initiate channel opening but were insufficient to allow K+ flow. Adjusting protonation states to represent the subsequent N520 intermediate in our MD simulations induced further conformational changes, including rearrangements of transmembrane helices 2 and 7, that opened the putative ion- translocation pathway. This allows spontaneous but low cation but not anion conduction, that matches experiments. Our findings provide critical structural insights into key intermediates of the channel opening mechanism, enhancing our understanding of ion conduction and selectivity in channelrhodopsins at an atomistic level.

biophysics↗

bioRxiv2022

Molecular snapshots of drug release from tubulin over eleven orders of magnitude in time

The binding and release of ligands from their protein targets is central to fundamental biological processes as well as to drug discovery. Photopharmacology introduces chemical triggers that allow the changing of ligand affinities and thus biological activity by light. Insight into the molecular mechanisms of photopharmacology is largely missing because the relevant transitions during the light-triggered reaction cannot be resolved by conventional structural biology. Using time-resolved serial crystallography at a synchrotron and X-ray free-electron laser, we captured the release of the anti-cancer compound azo-combretastatin A4 and the resulting conformational changes in tubulin. Nine structural snapshots from 1 ns to 100 ms complemented by simulations show how cis-to-trans isomerization of the azobenzene bond leads to a switch in ligand affinity, opening of an exit channel, and collapse of the binding pocket upon ligand release. The resulting global backbone rearrangements are related to the action mechanism of microtubule-destabilizing drugs.

molecular biology↗