Biology subjects

Neutze, R.

Publications and source records attributed to Neutze, R..

3 recordsLinked to original sources

bioRxiv2025

A user-friendly goniometer-compatible fixed-target platform formacromolecular crystallography at synchrotrons

Fixed-target platforms provide convenient support for microcrystals during serial X-ray crystallography studies using synchrotron radiation. Here, we describe a simple, user-friendly 3D-printed support where the crystals are sandwiched between two layers of thin X-ray transparent membrane resulting in very low scattering background. The platform is compatible with magnetic mounting onto the standard goniometer of macromolecular crystallography beamlines. Our design utilizes a 96-well frame that facilitates hanging-drop experiments directly on the membrane using conventional crystallization plates, thereby eliminating multiple pipetting and crystal handling steps. Crystals can be enclosed into a sandwich and packed into cassettes, preventing the risk of the sample drying out during room-temperature transportation to synchrotron sources. The versatility of the platform is demonstrated by five structures solved using different crystallization- and data-collection strategies. Single crystal rotational-crystallography at both room- and cryogenic-temperatures using large crystals of lysozyme is shown. On-chip microcrystallization is illustrated by use of a photosynthetic reaction center as an example. Finally, serial crystallography data collection at room-temperature from microcrystals of photosynthetic reaction center as well as cytochrome c oxidase crystallized in lipidic cubic phase is presented.

biophysics↗

bioRxiv2022

ULTRAFAST STRUCTURAL CHANGES DIRECT THE FIRST MOLECULAR EVENTS OF VISION

Vision is initiated by the rhodopsin family of light-sensitive G protein-coupled receptors (GPCRs). A photon is absorbed by the 11-cis retinal chromophore of rhodopsin which isomerises within 200 femtoseconds to the all-trans conformation, thereby initiating the cellular signal transduction processes that ultimately lead to vision. However, the intramolecular mechanism by which the photoactivated retinal induces the activation events inside rhodopsin remains elusive. In this work, we use ultrafast time-resolved crystallography at room temperature to determine how an isomerised twisted all-trans retinal stores the photon energy required to initiate protein conformational changes associated with the formation of the G protein-binding signalling state. The distorted retinal at 1 ps time-delay of photoactivation has pulled away from half of its numerous interactions with its binding pocket, and the excess of the photon energy is released through an anisotropic protein breathing motion in the direction of the extracellular space. Strikingly, the very early structural motions in the protein side chains of rhodopsin appear in regions involved in later stages of the conserved Class A GPCR activation mechanism. Our work sheds light on the earliest stages of vision in vertebrates and points to fundamental aspects of the molecular mechanisms of agonist-mediated GPCR activation.

biochemistry↗

bioRxiv2018

Nuclear envelope budding is an evolutionary conserved phenomenon

Nuclear envelope budding (NEB) is a recently discovered alternative pathway for nucleocytoplasmic communication distinct from the movement of material through the nuclear pore complex. Through quantitative electron microscopy and tomography, we demonstrate how NEB is evolutionarily conserved from early protists to human cells. In the yeast Saccharomyces cerevisiae, NEB events occur with higher frequency during heat shock, upon exposure to arsenite or hydrogen peroxide, and when the proteasome is inhibited. Yeast cells treated with azetidine-2-carboxylic acid, a proline analogue that induces protein misfolding, display the most dramatic increase in NEB, suggesting a causal link to protein quality control. This link was further supported by both localization of ubiquitin and Hsp104 to protein aggregates and NEB events, and the evolution of these structures during heat shock. We hypothesize that NEB is part of normal cellular physiology in a vast range of species and that in S. cerevisiae NEB comprises a stress response aiding the transport of protein aggregates across the nuclear envelope. Significance StatementA defining feature of eukaryotes is the nuclear envelope, a double lipid bilayer that serves to isolate and protect the cells genetic material. Transport of large molecules over this barrier is believed to occur almost exclusively via the nuclear pores. However, herpes virions and mega ribonucleoproteins (megaRNPs) use an alternative means of transport - via nuclear envelope budding (NEB). Here, we show NEB is a ubiquitous eukaryotic phenomenon and increases when exposed to various forms of cellular stress. NEB frequency was maximal when the cell was challenged with a drug that induces protein misfolding, indicating this transport pathway plays a role in protein quality control. These results imply that NEB is an underappreciated yet potentiallyfundamental means of nuclear transport.

cell biology↗