Biology subjects

Yorke, B. A.

Publications and source records attributed to Yorke, B. A..

4 recordsLinked to original sources

bioRxiv2026

Use of photocaged molecular oxygen for time-resolved serial X-ray crystallography

Photocages offer an attractive means of synchronously triggering enzyme-substrate driven reactions in biological systems in crystallo expanding the reach of light-driven catalysis. We describe the application of photocaged molecular oxygen to trigger molecular oxygen binding in crystals of myoglobin under anaerobic conditions and follow structural changes using both serial synchrotron and serial femtosecond X-ray crystallography. This is enabled through use of fixed targets under anaerobic conditions, utilising thin polymeric films with low molecular oxygen permeability and validated by serially collecting deoxy myoglobin structures and complementary in crystallo UV-Vis spectroscopy. Release of molecular oxygen from the photocage and subsequent binding of the gaseous ligand is structurally visualised in oxygen-bound structures of myoglobin at 5 and 10 ms and various laser parameters. We present a robust workflow for enabling anaerobic room-temperature data collection of oxygen-sensitive samples on fixed targets and report the successful photo-release of caged molecular oxygen for time-resolved serial crystallography.

molecular biology↗

bioRxiv2024

Adherence to the conservation of momentum to elucidate membrane transporter mechanisms.

1Diverse conformations of highly populated protein metastable states are well-studied, but the fleeting transitions between these states cannot be observed by experimental methods or molecular dynamics simulations. To address this, we present a computationally inexpensive algorithm, "cold-inbetweening", which generates trajectories in torsion angle space, by minimising the overall kinetic energy needed to complete a transition between experimentally determined end-states. Here we demonstrate the application of cold-inbetweening to provide mechanistic insight into the ubiquitous alternate access model of operation in three membrane transporter superfamilies. The model proposes mutually exclusive outward and inward pore opening, allowing ligand translocation but preventing damage from free solvent flow. Here, we study DraNramp from Deinococcus radiodurians, MalT from Bacillus cereus, and MATE from Pyrococcus furiosus. In MalT, the trajectory demonstrates elevator transport through unwinding of a supporter arm helix, maintaining adequate space to transport maltose. In DraNramp, outward-gate closure occurs prior to inward-gate opening, in agreement with the alternate access model. In the MATE transporter, switching conformation involves obligatory rewinding of an extended N-terminal helix to avoid steric backbone clashes. This concurrently plugs the cavernous ligand-binding site during the conformational change. More generally, cold-inbetweening can be used to inform hypotheses about large functionally relevant conformational changes.

biophysics↗

bioRxiv2023

An ultraviolet-driven rescue pathway for oxidative stress to eye lens protein human gamma-D crystallin.

1Human gamma-D crystallin (HGD) is the major constituent of the eye lens. Aggregation of HGD contributes to cataract formation, the leading cause of blindness worldwide. It is unique in its longevity, maintaining its folded and soluble state for 50-60 years. One outstanding question is the structural basis of this longevity despite oxidative aging and environmental stressors including ultraviolet radiation (UV). Here we present crystallographic structures evidencing a UV-induced crystallin redox switch mechanism. The room-temperature serial synchrotron crystallographic (SSX) structure of freshly prepared crystallin shows no post-translational modifications. After aging for nine months in the absence of light, a covalently bound reducing agent modifying surface cysteines is observed for the first time by low-dose SSX. This is shown to be UV-labile in an acutely light-exposed structure. This suggests a mechanism by which a major source of crystallin damage, UV, may also act as a rescuing factor in a finely balanced redox system.

biochemistry↗

bioRxiv2020

Diffusive dynamics of Aspartate α-decarboxylase (ADC) liganded with D-serine in aqueous solution

Incoherent neutron spectroscopy, in combination with dynamic light scattering was used to investigate the effect of ligand binding on the center-of-mass self-diffusion and internal diffusive dynamics of E.coli aspartate -decarboxylase (ADC). The X-ray crystal structure of the D-serine inhibitor complex with ADC was also determined, and molecular dynamics simulations used to further probe the structural rearrangements that occur as a result of ligand binding. These experiments reveal the existence of higher order oligomers of the ADC tetramer on ns-ms time-scales, and also show that ligand binding both affects the ADC internal diffusive dynamics and appears to further increase the size of the higher order oligomers.

biophysics↗