Biology subjects

Maia, F. R. N. C.

Publications and source records attributed to Maia, F. R. N. C..

4 recordsLinked to original sources

bioRxiv2024

Controlling Drug Partitioning in Individual Protein Condensates through Laser-Induced Microscale Phase Transitions.

Gelation of protein condensates formed by liquid-liquid phase separation (LLPS) occurs in a wide range of biological contexts, from the assembly of biomaterials to the formation of fibrillar aggregates and is therefore of interest for biomedical applications. Soluble-to-gel (sol-gel) transitions are controlled through macroscopic processes such as changes in temperature or buffer composition, resulting in bulk conversion of liquid droplets into microgels within minutes to hours. Using microscopy and mass spectrometry, we show that condensates of an engineered mini-spidroin (NT2repCTYF) undergo a spontaneous sol-gel transition resulting in the loss of exchange of proteins between the soluble and the condensed phase. We find that liquid spidroin condensates absorb visible light, which enables us to control sol-gel transitions of individual droplets through laser pulses. Fluorescence microscopy reveals that laser-induced gelation significantly alters the interactions between droplet proteins and small molecules, which allows us to load single droplets with an anticancer drug. In summary, our findings demonstrate direct control of phase transitions in individual condensates opening new avenues for functional and structural characterization. SYNOPSIS TOCThe liquid-to-solid transitions of phase-separated protein condensates are challenging to control. Leppert et al. show that condensates of engineered mini-spidroins gelate at slightly elevated temperatures. Using high-energy laser pulses at wavelengths that are absorbed by the droplets, the authors induce sol-gel transitions in single droplets. These gelated droplets are chemically stable and exhibit an increased ability to sequester drug molecules. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/584573v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@a7186corg.highwire.dtl.DTLVardef@345bd0org.highwire.dtl.DTLVardef@177c94forg.highwire.dtl.DTLVardef@1416456_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

Preparation and Characterization of Inactivated Tick-Borne Encephalitis Virus Samples for Single Particle Imaging at European XFEL

X-ray imaging of virus particles at European XFEL could eventually allow solving their complete structure, potentially approaching resolution of other structural virology methods. To achieve this ambitious goal with todays technologies, several mL of purified virus suspension containing at least 1012 particles per mL are required. Such large amounts of concentrated suspension have never before been obtained for enveloped viruses. Tick-borne encephalitis virus (TBEV) represents an attractive model system for the development of enveloped virus purification and concentration protocols, given the availability of large amounts of inactivated virus material provided by vaccine manufacturing facilities. Here we present the development of a TBEV vaccine purification and concentration scheme combined with a quality control protocol allowing substantial amounts of highly concentrated non-aggregated suspension to be obtained. Preliminary single particle imaging experiments were performed for this sample at European XFEL, showing distinct diffraction patterns.

biophysics↗

bioRxiv2018

Electrospray sample injection for single-particle imaging with X-ray lasers

The possibility of imaging single proteins constitutes an exciting challenge for X-ray lasers. Despite encouraging results on large particles, imaging small particles has proven to be difficult for two reasons: not quite high enough pulse intensity from currently available X-ray lasers and, as we demonstrate here, contamination of the aerosolised molecules by non-volatile contaminants in the solution. The amount of contamination on the sample depends on the initial droplet-size during aerosolisation. Here we show that with our electrospray injector we can decrease the size of aerosol droplets and demonstrate virtually contaminant-free sample delivery of organelles, small virions, and proteins. The results presented here, together with the increased performance of next generation X-ray lasers, constitute an important stepping stone towards the ultimate goal of protein structure determination from imaging at room temperature and high temporal resolution.

biophysics↗

bioRxiv2017

Cryo-EM Of A Marseilleviridae Virus Particle Reveals A Large Internal Microassembly

Nucleocytoplasmic large DNA viruses (NCLDVs) blur the line between viruses and cells. Melbournevirus (MelV, fam. Marseilleviridae) belongs to a new family of NCLDVs. Here we present an electron cryo-microscopy structure of the MelV particle, with the largest known triangulation number (T=309) for a virus. The 230-nm particle is constructed by 3080 pseudo-hexagonal capsomers and encloses a membrane bilayer. Its most distinct feature is a large dense body (LDB) consistently found in all particles. Electron cryo-tomography of 147 particles showed that the LDB is located preferentially in proximity to the bilayer. The LDB is 30 nm in size and its density matches that of a genome/protein complex. More than 58 proteins are associated with the purified particle, including histone-like proteins, putative membrane proteins and capsid proteins. The observed intricate structural organization reinforces the genetic complexity of MelV, setting it apart from other viruses, and suggests an evolutionary link with cellular organisms.

biophysics↗