Ageing drop by drop: Disturbance of the membrane-less organelle biogenesis as an aging hallmark.

Despite extensive research, the features associated with the aging phenotype are not all-inclusive and need to be updated on a regular basis to incorporate new findings. We propose to include the dysfunction of membrane-less organelle (MLO) as a new aging hallmark. Special scaffold proteins with a high degree of intrinsic disorder drive the formation of MLOs via the liquid-liquid phase separation (LLPS) process.

Crystallographic insights into lipid-membrane protein interactions in microbial rhodopsins.

The primary goal of our work is to provide structural insights into the influence of the hydrophobic lipid environment on the membrane proteins (MPs) structure and function. Our work will not cover the well-studied hydrophobic mismatch between the lipid bilayer and MPs. Instead, we will focus on the less-studied direct molecular interactions of lipids with the hydrophobic surfaces of MPs.

Production of eukaryotic heliorhodopsins for structural analysis utilizing the LEXSY expression system.

Heliorhodopsins (HeRs) constitute a novel and distinct group of microbial rhodopsins, characterized by the inverted position of C- and N- termini relative to conventional Type I rhodopsins. The production of HeRs for structural and functional investigations has proven challenging, as evidenced by the structural elucidation of only two proteins and the functional characterization of a few others to date. Notably, no eukaryotic HeRs have been reported thus far.

Examining transfer of TERT to mitochondria under oxidative stress.

The primary role of telomerase is the lengthening of telomeres. Nonetheless, emerging evidence highlights additional functions of telomerase outside of the nucleus. Specifically, its catalytic subunit, TERT (Telomerase Reverse Transcriptase), is detected in the cytosol and mitochondria.

Calcium-Sensitive Microbial Rhodopsin VirChR1: A Femtosecond to Second Photocycle Study.

Viral rhodopsins are light-gated cation channels representing a novel class of microbial rhodopsins. For viral rhodopsin 1 subfamily members VirChR1 and OLPVR1, channel activity is abolished above a certain calcium concentration. Here we present a calcium-dependent spectroscopic analysis of VirChR1 on the femtosecond to second time scale.

A subgroup of light-driven sodium pumps with an additional Schiff base counterion.

Light-driven sodium pumps (NaRs) are unique ion-transporting microbial rhodopsins. The major group of NaRs is characterized by an NDQ motif and has two aspartic acid residues in the central region essential for sodium transport. Here we identify a subgroup of the NDQ rhodopsins bearing an additional glutamic acid residue in the close vicinity to the retinal Schiff base.

Order wrapped in chaos: On the roles of intrinsically disordered proteins and RNAs in the arrangement of the mitochondrial enzymatic machines.

The analysis of cryo-electron tomography images of human and rat mitochondria revealed that the mitochondrial matrix is at least as crowded as the cytosol. To mitigate the crowding effects, metabolite transport in the mitochondria primarily occurs through the intermembrane space, which is significantly less crowded. The scientific literature largely ignores how enzyme systems and metabolite transport are organized in the crowded environment of the mitochondrial matrix.

Intracellular microbial rhodopsin-based optogenetics to control metabolism and cell signaling.

Microbial rhodopsin (MRs) ion channels and pumps have become invaluable optogenetic tools for neuroscience as well as biomedical applications. Recently, MR-optogenetics expanded towards subcellular organelles opening principally new opportunities in optogenetic control of intracellular metabolism and signaling precise manipulations of organelle ion gradients using light. This new optogenetic field expands the opportunities for basic and medical studies of cancer, cardiovascular, and metabolic disorders, providing more detailed and accurate control of cell physiology.

Channelrhodopsin-2 Oligomerization in Cell Membrane Revealed by Photo-Activated Localization Microscopy.

Microbial rhodopsins are retinal membrane proteins that found a broad application in optogenetics. The oligomeric state of rhodopsins is important for their functionality and stability. Of particular interest is the oligomeric state in the cellular native membrane environment.

Hijacking of internal calcium dynamics by intracellularly residing viral rhodopsins.

Rhodopsins are ubiquitous light-driven membrane proteins with diverse functions, including ion transport. Widely distributed, they are also coded in the genomes of giant viruses infecting phytoplankton where their function is not settled. Here, we examine the properties of OLPVR1 (Organic Lake Phycodnavirus Rhodopsin) and two other type 1 viral channelrhodopsins (VCR1s), and demonstrate that VCR1s accumulate exclusively intracellularly, and, upon illumination, induce calcium release from intracellular IP-dependent stores.

Light-Driven Sodium Pump as a Potential Tool for the Control of Seizures in Epilepsy.

The marine flavobacterium Krokinobactereikastus light-driven sodium pump (KR2) generates an outward sodium ion current under 530 nm light stimulation, representing a promising optogenetic tool for seizure control. However, the specifics of KR2 application to suppress epileptic activity have not yet been addressed. In the present study, we investigated the possibility of KR2 photostimulation to suppress epileptiform activity in mouse brain slices using the 4-aminopyrindine (4-AP) model.

Retinal-Based Anion Pump from the Cyanobacterium Tolypothrix campylonemoides.

In this work, TcaR rhodopsin from the cyanobacterium Tolypothrix campylonemoides was characterized. Analysis of the amino acid sequence of TcaR revealed that this protein possesses a TSD motif that differs by only one amino acid from the TSA motif of the known halorhodopsin chloride pump. The TcaR protein was expressed in E.

The TR-icOS setup at the ESRF: time-resolved microsecond UV-Vis absorption spectroscopy on protein crystals.

The technique of time-resolved macromolecular crystallography (TR-MX) has recently been rejuvenated at synchrotrons, resulting in the design of dedicated beamlines. Using pump-probe schemes, this should make the mechanistic study of photoactive proteins and other suitable systems possible with time resolutions down to microseconds. In order to identify relevant time delays, time-resolved spectroscopic experiments directly performed on protein crystals are often desirable.

Mechanisms of inward transmembrane proton translocation.

Proton transport is indispensable for cell life. It is believed that molecular mechanisms of proton movement through different types of proton-conducting molecules have general universal features. However, elucidation of such mechanisms is a challenge.

Flotillin-associated rhodopsin (FArhodopsin), a widespread paralog of proteorhodopsin in aquatic bacteria with streamlined genomes.

Microbial rhodopsins are found more than once in a single genome (paralogs) often have different functions. We screened a large dataset of open ocean single-amplified genomes (SAGs) for co-occurrences of multiple rhodopsin genes. Many such cases were found among Pelagibacterales (SAR11), HIMB59, and the Gammaproteobacteria SAGs.

Mirror proteorhodopsins.

Proteorhodopsins (PRs), bacterial light-driven outward proton pumps comprise the first discovered and largest family of rhodopsins, they play a significant role in life on the Earth. A big remaining mystery was that up-to-date there was no described bacterial rhodopsins pumping protons at acidic pH despite the fact that bacteria live in different pH environment. Here we describe conceptually new bacterial rhodopsins which are operating as outward proton pumps at acidic pH.

Structural insights into the effects of glycerol on ligand binding to cytochrome P450.

New antitubercular drugs are vital due to the spread of resistant strains. Carbethoxyhexyl imidazole (CHImi) inhibits cytochrome P450 CYP124, which is a steroid-metabolizing enzyme that is important for the survival of Mycobacterium tuberculosis in macrophages. The available crystal structure of the CYP124-CHImi complex reveals two glycerol molecules in the active site.

Crystal structure of semi-synthetic obelin-v after calcium induced bioluminescence implies coelenteramine as the main reaction product.

Coelenterazine-v (CTZ-v), a synthetic vinylene-bridged π-extended derivative, is able to significantly alter bioluminescence spectra of different CTZ-dependent luciferases and photoproteins by shifting them towards longer wavelengths. However, Ca-regulated photoproteins activated with CTZ-v display very low bioluminescence activities that hampers its usage as a substrate of photoprotein bioluminescence. Here, we report the crystal structure of semi-synthetic Ca-discharged obelin-v bound with the reaction product determined at 2.

Structural insights into light-driven anion pumping in cyanobacteria.

Transmembrane ion transport is a key process in living cells. Active transport of ions is carried out by various ion transporters including microbial rhodopsins (MRs). MRs perform diverse functions such as active and passive ion transport, photo-sensing, and others.

Ferritin self-assembly, structure, function, and biotechnological applications.

Ferritin is a vital protein complex responsible for storing iron in almost all living organisms. It plays a crucial role in various metabolic pathways, inflammation processes, stress response, and pathogenesis of cancer and neurodegenerative diseases. In this review we discuss the role of ferritin in diseases, cellular iron regulation, its structural features, and its role in biotechnology.

Development and Characterization of Flavin-Binding Fluorescent Proteins, Part II: Advanced Characterization.

Flavin-based fluorescent proteins (FbFPs), a class of small fluorescent proteins derived from light-oxygen-voltage (LOV) domains, bind ubiquitous endogenous flavins as chromophores. Due to their unique properties, they can be used as versatile in vivo reporter proteins under aerobic and anaerobic conditions. This chapter presents methodologies for in-depth characterization of the biochemical, spectroscopic, photophysical, and photochemical properties of FbFPs.