Biology subjects

Myasnikov, A. G.

Publications and source records attributed to Myasnikov, A. G..

5 recordsLinked to original sources

bioRxiv2022

Inside and outside of virus-like particles HBc and HBc/4M2e: a comprehensive study of the structure

The structure of the core particles of the hepatitis B virus core antigen (HBc) with the insertion of four external domains of the influenza A M2 protein (HBc/4M2e) have been studied using a combination of molecular modeling and cryogenic electron microscopy (cryo-EM). It has also been shown that synthesis of particles occurs inside bacterial cells, but despite the big inner volume of the core shell particle, purified HBc/4M2e do not contain detectable amounts of bacterial proteins. It has been shown that a fragment of the M2e is prone to the formation of amyloid-like fibrils. However, as a part of HBc immunodominant loop M2e domains as repeats, does not exhibit a tendency to aggregation. Full-atom HBc-4M2E model with 3A resolution was obtained by molecular modeling methods based on cryo-EM data.

biophysics↗

bioRxiv2021

Directed evolution of the rRNA methylating enzyme Cfr reveals molecular basis of antibiotic resistance

Alteration of antibiotic binding sites through modification of ribosomal RNA (rRNA) is a common form of resistance to ribosome-targeting antibiotics. The rRNA-modifying enzyme Cfr methylates an adenosine nucleotide within the peptidyl transferase center, resulting in the C-8 methylation of A2503 (m8A2503). Acquisition of cfr results in resistance to eight classes of ribosome-targeting antibiotics. Despite the prevalence of this resistance mechanism, it is poorly understood whether and how bacteria modulate Cfr methylation to adapt to antibiotic pressure. Moreover, direct evidence for how m8A2503 alters antibiotic binding sites within the ribosome is lacking. In this study, we performed directed evolution of Cfr under antibiotic selection to generate Cfr variants that confer increased resistance by enhancing methylation of A2503 in cells. Increased rRNA methylation is achieved by improved expression and stability of Cfr through transcriptional and post-transcriptional mechanisms, which may be exploited by pathogens under antibiotic stress as suggested by natural isolates. Using a variant which achieves near-stoichiometric methylation of rRNA, we determined a 2.2 [A] cryo-EM structure of the Cfr-modified ribosome. Our structure reveals the molecular basis for broad resistance to antibiotics and will inform the design of new antibiotics that overcome resistance mediated by Cfr.

biochemistry↗

bioRxiv2020

Cryo-EM structures reveal a multistep mechanism of Hsp90 activation by co-chaperone Aha1

Hsp90 is a ubiquitous molecular chaperone that facilitates the folding and maturation of hundreds of cellular “client” proteins. The ATP-driven client maturation process is regulated by a large number of co-chaperones. Among them, Aha1 is the most potent activator of Hsp90 ATPase activity and thus dramatically affects Hsp90’s client proteins. To understand the Aha1 activation mechanism, we determined full-length Hsp90:Aha1 structures in six different states by cryo-electron microscopy, including nucleotide-free semi-closed, nucleotide-bound pre-hydrolysis, and semi-hydrolyzed states. Our structures demonstrate that the two Aha1 domains can each interact with Hsp90 in two different modes, uncovering a complex multistep activation mechanism. The results show that Aha1 accelerates the chemical step of ATP hydrolysis like a conventional enzyme, but most unusually, catalyzes the rate-limiting large-scale conformational changes of Hsp90 fundamentally required for ATP hydrolysis. Our work provides a structural framework to guide small molecule development targeting this critical modulator of client protein maturation.Competing Interest StatementThe authors have declared no competing interest.View Full Text

biophysics↗

bioRxiv2019

Allosteric coupling between alpha-rings of the 20S proteasome

The proteasomal machinery performs essential regulated protein degradation in eukaryotes. Classic proteasomes are symmetric, with a regulatory ATPase docked at each end of the cylindrical 20S. Asymmetric complexes are also present in cells, either with a single ATPase or with an ATPase and non-ATPase at two opposite ends. The mechanism that populates these different proteasomal complexes is unknown. Using archaea homologs, we constructed asymmetric forms of proteasomes. We demonstrate that the gate conformation of two opposite ends of 20S are coupled: binding one ATPase opens a gate locally, and also the remote opposite gate allosterically. Such allosteric coupling leads to cooperative binding of proteasomal ATPases to 20S, and promotes formation of proteasomes symmetrically configured with two identical ATPases. It may also promote formation of asymmetric complexes with an ATPase and a non-ATPase at opposite ends. We propose that in eukaryotes a similar mechanism regulates the composition of the proteasomal population.

biochemistry↗

bioRxiv2018

Structural basis of eIF2B-catalyzed GDP exchange and phosphoregulation by the integrated stress response

The integrated stress response (ISR) tunes the rate of protein synthesis. Control is exerted by phosphorylation of the general translation initiation factor eIF2. eIF2 is a GTPase, that becomes activated by eIF2B, a two-fold symmetric and heterodecameric complex that functions as eIF2s dedicated nucleotide exchange factor. Phosphorylation converts eIF2 from a substrate into an inhibitor of eIF2B. We report cryoEM structures of eIF2 bound to eIF2B in the dephosphorylated state. The structures reveal that the eIF2B decamer is a static platform upon which one or two flexible eIF2 trimers bind and align with eIF2Bs bipartite catalytic centers to catalyze guanine nucleotide exchange. Phosphorylation refolds eIF2, allowing it to contact eIF2B at a different interface and, we surmise, thereby sequesters it into a non-productive complex.\n\nOne Sentence SummaryStructures of translation factors eIF2 and eIF2B reveal the mechanism of nucleotide exchange and its phosphoregulation during stress.

biochemistry↗