Biology subjects

Abdelhamid, M. A. S.

Publications and source records attributed to Abdelhamid, M. A. S..

3 recordsLinked to original sources

bioRxiv2025

A new protein-dependent riboswitch activates ribosomal frameshifting

Programmed -1 ribosomal frameshifting (PRF) is a translational control mechanism used by RNA viruses to regulate the relative abundance of proteins encoded in different reading frames. Cardioviruses exhibit the highest known PRF efficiency, with ~85% of ribosomes shifting into the -1 frame. This unusual event requires an interaction between the viral 2A protein and a stimulatory element in the RNA genome, but the basis for protein-dependence is unclear. To address this, here we investigate structure and dynamics of the PRF signal in Theiler's murine encephalitis virus (TMEV). By combining X-ray crystallography, small angle X-ray scattering (SAXS) and single-molecule fluorescence resonance energy transfer (smFRET), we show that 2A binding switches the RNA from a stem-loop conformation into a pseudoknot, and we demonstrate that pseudoknot formation is essential for efficient PRF in vitro and in cells. Together, these findings illustrate how the cardiovirus PRF element behaves as a protein-dependent riboswitch, defining the molecular mechanism by which frameshifting is conditionally activated.

molecular biology↗

bioRxiv2024

Kinesin-1 is highly flexible and adopts an open conformation in the absence of cargo

Kinesin-1 is an essential anterograde microtubule motor protein. The core kinesin motor is a homodimer of two heavy chains; N-terminal motor domains hydrolyse ATP and walk along microtubules, whilst a long elongated coiled-coil stalk and an intrinsically disordered C-terminal tail region bind cargos. Kinesin autoinhibition is key to preventing futile ATP consumption and occurs, at least in part, through direct interactions between N-terminal motor domains and C-terminal inhibitory motifs. Despite significant advances in our understanding of kinesin walking, little is known about the kinesin-1 conformational landscape of the stalk and tail domains. Here we apply solution based biophysical analysis tools to study conformational changes in kinesin-1, with full rotational freedom, and in response to changes in ionic strength, mutations, and the presence of microtubules. This has allowed us to uncover the inherent flexibility in kinesin-1 which gives insights into autoinhibition and the regulation of intracellular transport.

biophysics↗

bioRxiv2024

Rep structures can be tuned by ionicity via metastable intermediates in the absence of DNA

DNA helicases undergo conformational changes; however, their structural dynamics are poorly understood. Here, we study single molecules of superfamily 1A DNA helicase Rep, which undergo conformational transitions during bacterial DNA replication, repair and recombination. We use time-correlated single-photon counting (TCSPC), fluorescence correlation spectroscopy (FCS), rapid single-molecule Forster resonance energy transfer (smFRET), Anti-Brownian ELectrokinetic (ABEL) trapping and molecular dynamics simulations (MDS) to provide unparalleled temporal and spatial resolution of Reps domain movements. We detect four states revealing two hitherto hidden intermediates (S2, S3), between the open (S1) and closed (S4) structures, whose stability is salt dependent. Reps open-to-closed switch involves multiple changes to all four subdomains 1A, 1B, 2A and 2B along the S1[->]S2[->]S3[->]S4 transitional pathway comprising an initial truncated swing of 2B which then rolls across the 1B surface, following by combined rotations of 1B, 2A and 2B. High forward and reverse rates for S1[->]S2 suggest that 1B may act to frustrate 2B movement to prevent premature Rep closure in the absence of DNA. These observations support a more general binding model for accessory DNA helicases that utilises conformational plasticity to explore a multiplicity of structures whose landscape can be tuned by salt prior to locking-in upon DNA binding.

biophysics↗