Biology subjects

Tzokov, S. B.

Publications and source records attributed to Tzokov, S. B..

4 recordsLinked to original sources

bioRxiv2024

Molecular determinants of protein pathogenicity at the single-aggregate level

Determining the structure-function relationships of protein aggregates is a fundamental challenge in biology. These aggregates, whether formed in vitro, within cells, or in living organisms, present significant heterogeneity in their molecular features such as size, structure, and composition, making it difficult to determine how their structure influences their functions. Interpreting how these molecular features translate into functional roles is crucial for understanding cellular homeostasis and the pathogenesis of various debilitating diseases like Alzheimers and Parkinsons. In this study, we introduce a bottom-up approach to explore how variations in protein aggregates size, composition, post-translational modifications and point mutations profoundly influence their biological functions. Applying this method to Alzheimers and Parkinsons associated proteins, we uncover the mechanism of novel disease-relevant pathways and demonstrate how subtle alterations in composition and morphology can shift the balance between healthy and pathological states. Our findings establish a broadly applicable framework for investigating protein dysfunctions in various proteinopathies.

biochemistry↗

bioRxiv2024

CryoEM Reconstruction of Yeast ADP-Actin Filament at 2.5 Angstrom resolution. A comparison with mammalian and avian F-actin.

The core component of the actin cytoskeleton is the globular protein G-actin, which reversibly polymerises into filaments (F-actin). Budding yeast possesses a single actin which shares 87-89% sequence identity with vertebrate actin isoforms. Previous structural studies indicate very close overlap of main-chain backbones. Intriguingly however, substitution of yeast ACT1 with vertebrate {beta}-cytoplasmic actin severely disrupts cell function and substitution with a skeletal muscle isoform is lethal. Here we report a 2.5 [A] structure of budding yeast F-actin. Previously unresolved side-chain information now highlights four main differences in the comparison of yeast and vertebrate ADP F-actins: a more open nucleotide binding pocket; a more solvent exposed C-terminus; a rearrangement of intersubunit binding interactions in the vicinity of the D-loop and changes in the hydrogen bonding network in the vicinity of histidine 73 (yeast actin) and methyl-histidine 73 (vertebrate actin).

biochemistry↗

bioRxiv2021

The cryo-EM structure of the bacterial type I DNA segregation ATPase filament reveals its conformational plasticity upon DNA binding.

The efficient segregation of replicated genetic material is an essential step for cell division. In eukaryotic cells, sister chromatids are separated via the mitotic spindles. In contrast, bacterial cells use several evolutionarily-distinct genome segregation systems. The most common of these is the Type I Par system. It consists of an adapter protein, ParB, that binds to the DNA cargo via interaction with the parS DNA sequence; and an ATPase, ParA, that binds nonspecific DNA and mediates cargo transport. However, the molecular details of how this system functions are not well understood. Here, we report the cryo-EM structure of a ParA filament bound to its DNA template, using the chromosome 2 (Chr2) of Vibrio cholerae as a model system. We also report the crystal structures of this protein in various nucleotide states, which collectively offer insight into its conformational changes from dimerization through to DNA binding and filament assembly. Specifically, we show that the ParA dimer is stabilized by nucleotide binding, and forms a left-handed filament using DNA as a scaffold. Our structural analyses also reveal dramatic structural rearrangements upon DNA binding and filament assembly. Finally, we show that filament formation is controlled by nucleotide hydrolysis. Collectively, our data provide the structural basis for ParAs cooperative binding to DNA and the formation of high ParA density regions on the nucleoid, and suggest a role for its filament formation.

biophysics↗

bioRxiv2019

The cryo-EM structure of the bacterial flagellum cap complex suggests a molecular mechanism for filament elongation.

The bacterial flagellum is a remarkable molecular motor, present at the surface of many bacteria, whose primary function is to allow motility through the rotation of a long filament protruding from the bacterial cell. A cap complex, consisting of an oligomeric assembly of the protein FliD, is localized at the tip of the flagellum, and is essential for filament assembly, as well as adherence to surfaces in some bacteria. However, the structure of the intact cap complex, and the molecular basis for its interaction with the filament, remains elusive. Here we report the cryo-EM structure of the Campylobacter jejuni cap complex. This structure reveals that FliD is pentameric, with the N-terminal region of the protomer forming an unexpected extensive set of contacts across several subunits, that contribute to FliD oligomerization. We also demonstrate that the native C. jejuni flagellum filament is 11-stranded and propose a molecular model for the filament-cap interaction.

microbiology↗