Biology subjects

Weik, M.

Publications and source records attributed to Weik, M..

5 recordsLinked to original sources

bioRxiv2026

Structural basis of the two-photon photoactivation mechanism of orange carotenoid protein

Cyanobacteria have produced Earths oxygen for 2.4 billion years by adapting to fluctuating irradiance. This adaptation relies on orange carotenoid protein (OCP), which mediates light-intensity- dependent photoprotective energy dissipation using a unique two-photon absorption mechanism. Photon absorption by ground-state OCP (OCPO) generates a metastable intermediate (OCP1h{nu}) that either relaxes thermally or, upon absorption of a second photon within [~]1 s, converts to the active photoprotective state (OCPR). By integrating static and time-resolved crystallography, cryo-EM, computation, spectroscopy and biochemistry, we assign the structure of OCP1h{nu}, establish its functional relevance and capture structural snapshots along the OCPO[->]OCP1h{nu} and OCP1h{nu}[->]OCPR photochemical pathways. We elucidate the molecular mechanism of OCP, which serves as a unique biological circuit breaker protecting the photosynthetic machinery from high light flux.

biochemistry↗

bioRxiv2024

Crystal structure of a bacterial photoactivated adenylate cyclase determined at room temperature by serial femtosecond crystallography

OaPAC is a recently discovered blue-light using flavin adenosine dinucleotide (BLUF) photoactivated adenylate cyclase from the cyanobacterium Oscillatoria acuminata that uses adenosine triphosphate and translates the light signal into the production of cyclic adenosine monophosphate. Here, we report the crystal structures of the enzyme in the absence of its natural substrate determined from room temperature serial crystallography data collected at both an X-ray free electron laser and a synchrotron and we compare them with the cryo macromolecular crystallography structures obtained at a synchrotron by us and others. These results reveal slight differences in the structure of the enzyme due to data collection at different temperatures and X-ray sources. We further investigate the effect of the Y6 mutation in the blue-light using flavin adenosine dinucleotide domain, a mutation which results in a rearrangement of the hydrogen-bond network around the flavin and a notable rotation of the side-chain of the critical Q48 residue. These studies pave the way for ps - ms time-resolved serial crystallography experiments at X-ray free electron lasers and synchrotrons in order to determine the early structural intermediates and correlate them with the well-studied ps - ms spectroscopic intermediates. SynopsisStructures of the dark-adapted state of a photoactivated adenylate cyclase are determined from serial crystallography (SX) data collected at room temperature at an X-ray free electron laser (XFEL) and a synchrotron and are compared with cryo macromolecular crystallography (MX) synchrotron structures obtained by us and others. These structures of the wild-type enzyme in combination with the cryo MX synchrotron structure of a light-sensor domain mutant provide insight into the hydrogen bond network rearrangement upon blue-light illumination and pave the way for the determination of structural intermediates of the enzyme by time-resolved SX.

biophysics↗

bioRxiv2023

Mutations in tau protein promote aggregation by favoring extended conformations

Amyloid aggregation of the intrinsically disordered protein (IDP) tau is involved in several diseases, called tauopathies. Mutations in the gene encoding tau are responsible for a class of inherited tauopathies called frontotemporal dementia and parkinsonism linked to chromosome 17Q (FTDP-17). These mutations are thought to trigger FTDP-17 by favoring the formation of tau amyloid fibrils. This work aims at deciphering the mechanisms through which the diseases-associated single point mutations promote amyloid formation. We combined biochemical characterization and small angle X-ray scattering (SAXS) to study six different FTDP-17 derived mutations. We found that the mutations promote aggregation to different degrees and can modulate tau conformational ensembles, intermolecular interactions and liquid-liquid phase separation propensity. In particular, we found a good correlation between the aggregation lag time of the mutants and their radius of gyration. We show that mutations disfavor intramolecular protein interactions which in turn favor extended conformations and promote amyloid aggregation. This work proposes a new connection between the structural features of tau monomers and their propensity to aggregate, providing a novel assay to evaluate aggregation propensity of tau variants.

biophysics↗

bioRxiv2022

Influence of pump laser fluence on ultrafast structural changes in myoglobin

High-intensity femtosecond pulses from an X-ray free-electron laser enable pump probe experiments for investigating electronic and nuclear changes during light-induced reactions. On time scales ranging from femtoseconds to milliseconds and for a variety of biological systems, time-resolved serial femtosecond crystallography (TR-SFX) has provided detailed structural data for light-induced isomerization, breakage or formation of chemical bonds and electron transfer1. However, all ultra-fast TR-SFX studies to date have employed such high pump laser energies that several photons were nominally absorbed per chromophore2-14. As multiphoton absorption may force the protein response into nonphysiological pathways, it is of great concern15 whether this experimental approach16 allows valid inferences to be drawn vis-a-vis biologically relevant single-photon-induced reactions17. Here we describe ultrafast pump-probe SFX experiments on photodissociation of carboxymyoglobin, showing that different pump laser fluences yield markedly different results. In particular, the dynamics of structural changes and observed indicators of the mechanistically important coherent oscillations of the Fe-CO bond distance (predicted by recent quantum wavepacket dynamics15) are seen to depend strongly on pump laser energy. Our results confirm both the feasibility and necessity of performing TR-SFX pump probe experiments in the linear photoexcitation regime. We consider this to be a starting point for reassessing design and interpretation of ultrafast TR-SFX pump probe experiments16 such that biologically relevant insight emerges.

biophysics↗

bioRxiv2020

Zinc Determines Dynamical Properties and Aggregation Kinetics of Human Insulin

Protein aggregation is a widespread process leading to deleterious consequences in the organism, with amyloid aggregates being important not only in biology but also for drug design and biomaterial production. Insulin is a protein largely used in diabetes treatment and its amyloid aggregation is at the basis of the so-called insulin-derived amyloidosis. Here we uncover the major role of zinc in both insulin dynamics and aggregation kinetics at low pH, where the formation of different amyloid superstructures (fibrils and spherulites) can be thermally induced. Amyloid aggregation is accompanied by zinc release and the suppression of water-sustained insulin dynamics, as shown by particle-induced X-ray emission and X-ray absorption spectroscopy and by neutron spectroscopy, respectively. Our study shows that zinc binding stabilizes the native form of insulin by facilitating hydration of this hydrophobic protein and suggests that introducing new binding sites for zinc can improve insulin stability and tune its aggregation propensity.Statement of Significance Localized amyloidosis occurs at insulin injection sites for diabetes treatment, leading to deleterious inflammations known as insulin-derived amyloidosis. Amyloid superstructures are also promising candidates in the field of biomaterials. Here we revealed that zinc, coordinated to insulin in the native form, is released upon amyloid aggregation, when insulin forms superstructures known as fibrils and spherulites. Zinc release leads to a full suppression of functionally essential protein dynamics through a modification of the protein’s hydration properties and completely modifies insulin amyloid kinetics. The results suggest that changes in protein hydration upon zinc binding/release modifies both stability and dynamics of insulin and might then be a general strategy to control protein stability and tune protein aggregation into amorphous and ordered superstructures.View Full Text

biophysics↗