Biology subjects

Orekhov, P. S.

Publications and source records attributed to Orekhov, P. S..

3 recordsLinked to original sources

bioRxiv2026

Structural insights into human recoverin

Recoverin is a key calcium sensor that controls the desensitization of the visual rhodopsin by GRK1. Previous studies have traditionally been conducted on bovine protein (bRec), while data on human ortholog (hRec) remain scarce. Here, we combine X-ray crystallography, X-ray absorption spectroscopy (XANES), quantum mechanical calculations, molecular dynamics, and functional assays to provide an integrated characterization of hRec. The 2Ca2+-bound hRec structure was solved at 1.60 [A], showing that, unlike bRec, hRec interacts with ROS membranes at physiologically relevant submicromolar Ca2+ levels, due to a species-specific charge distribution that might influence membrane interactions. Both recoverins form a set of Ca2+/Zn2+-bound conformers with improved functional performance. X-ray crystallography (1.85 [A]) and XANES revealed a specific tetrahedral Zn2+ site in 1Ca2+-bound hRec, the first such site reported in the NCS family. In 1Ca2+-bound hRec, zinc promotes the formation of active state, whereas in 2Ca2+-state of bRec, it significantly enhances GRK1 binding, as the latter can complement the Zn2+ coordination. These data refine our understanding of recoverin function in humans and highlight its role as a key link between calcium and zinc signaling in mammalian photoreceptors under normal and pathological conditions.

biochemistry↗

bioRxiv2025

Control of the force-bearing properties of microtubule-associated proteins via stalk/linker regions: insights from the NDC80 complex

Many microtubule-associated proteins (MAPs) function under mechanical loads. Among them, motor proteins and passive couplers link microtubules with other cytoskeletal filaments, membranous structures and diverse scaffolds to enable cell shape changes, locomotion and other important processes. A key kinetochore complex, NDC80, transmits forces from microtubule disassembly to chromosome motion during cell division. Recently, this complex has been shown to detach from microtubules more easily when pulled toward the minus-end of the microtubule than when pulled in the plus-end direction. Here, we used coarse-grained molecular dynamics and Brownian dynamics simulations to explain the asymmetric effect of the directional load on the unbinding of the NDC80 complex from microtubules and then generalized our findings to other MAPs. We found that the lever arm created by the stiff stalk of NDC80 tilted toward the plus-end of the microtubule is critical for asymmetric unbinding of this complex, similar to that of dynein. In contrast, EB-proteins, the microtubule crosslinker PRC1, and kinesins are predicted to lack pronounced unbinding asymmetry, either due to their almost perpendicular anchorage to the microtubule wall or due to the high flexibility of their linker regions proximal to the microtubule-binding domains. Thus, our study highlights some of the design principles of MAPs, explaining how their distal parts can impart, modulate or eliminate the dependence of unbinding on the direction of external loads. This information deepens our understanding of the load-bearing properties and functions of diverse MAPs and may guide the design of synthetic protein systems with predefined mechanical characteristics.

biophysics↗

bioRxiv2021

Effects of Mutations in the Receptor-Binding Domain of SARS-CoV-2 Spike on its Binding Affinity to ACE2 and Neutralizing Antibodies Revealed by Computational Analysis

SARS-CoV-2 causing coronavirus disease 2019 (COVID-19) is responsible for one of the most deleterious pandemics of our time. The interaction between the ACE2 receptors at the surface of human cells and the viral Spike (S) protein triggers the infection making the receptor-binding domain (RBD) of the SARS-CoV-2 S-protein a focal target for the neutralizing antibodies (Abs). Despite the recent progress in the development and deployment of vaccines, the emergence of novel variants of SARS-CoV-2 insensitive to Abs produced in response to the vaccine administration and/or monoclonal ones represents upcoming jeopardy. Here, we assessed the possible effects of single and multiple mutations in the RBD of SARS-CoV-2 S-protein on its binding energy to various antibodies and the human ACE2 receptor. The performed computational analysis indicates that while single amino acid replacements in RBD may only cause partial impairment of the Abs binding, moreover, limited to specific epitopes, some variants of SARS-CoV-2 (with as few as 8 mutations), which are already present in the population, may potentially result in a much broader antigenic escape. We also identified a number of point mutations, which, in contrast to the majority of replacements, reduce RBD affinity to various antibodies without affecting its binding to ACE2. Overall, the results provide guidelines for further experimental studies aiming at the identification of the high-risk RBD mutations allowing for an antigenic escape.

biophysics↗