Biology subjects

Hossain, K. A.

Publications and source records attributed to Hossain, K. A..

4 recordsLinked to original sources

bioRxiv2026

Structural basis of selective phospholipid recognition and local membrane deformation by CTL1

The physiological function of the human choline transporter-like protein CTL1 (SLC44A1) remains elusive despite its implication in key pathophysiological processes. Here we combine cryo-electron microscopy, native mass spectrometry, lipidomics, molecular dynamics simulations and functional assays to characterise the molecular basis of phospholipid recognition by CTL1 and explore its potential molecular role. High-resolution structures of CTL1 in both monomeric and dimeric states reveal a broad amphipathic cavity and an extracellular domain that accommodate multiple phospholipid species, identifying phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol and phosphatidic acid as endogenous CTL1 ligands. Molecular dynamics simulations show spontaneous phospholipid recruitment into these conserved binding sites accompanied by membrane thinning, curvature and local phospholipid reorganisation, with membrane deformation being enhanced upon dimerisation. Functional analyses in mammalian cells and in reconstituted proteoliposomes failed to detect CTL1-mediated choline transport, supporting a function distinct from canonical choline uptake. Together, our findings suggest that CTL1 acts as a phospholipid-binding protein capable of locally deforming the membrane and provide a structural framework linking phospholipid recognition to membrane organisation. These results offer new insights into potential mechanisms underlying the altered phospholipid homeostasis associated with CTL1 dysfunction in neurological diseases and cancer.

biophysics↗

bioRxiv2026

When lipids embrace RNA: pH-driven dynamics and mechanisms of LNP-mediated siRNA delivery

We present a comprehensive analysis of an efficient lipid nanoparticle (LNP) formulation that exhibits strong nucleic-acid delivery and potent inhibition of targeted RNAs. Using a combination of coarse-grained and atomistic molecular dynamics simulations, we characterized the pH-dependent structure of both unloaded and RNA-loaded LNPs, elucidated the mechanism of RNA encapsulation, and used these models to propose a plausible mechanism of endosomal escape. Consistent with prior experimental and computational studies, our simulations reproduce an inverted-hexagonal-type morphology in RNA-loaded LNPs, in which a hydrated core provides a polar environment suitable for accommodating RNA, whose charge is neutralized mainly by ionizable lipids that remain protonated near the RNA even at high pH, thereby bridging the RNA with the surrounding lipid environment. This structural picture is consistently observed across our multiscale simulations, with smaller self-assembled LNP mimetics reproducing the same local organization at both coarse-grained and atomistic resolution. A potential mechanism of endosomal escape emerges spontaneously from the simulations, involving stalk formation between the LNP and the endosomal membrane, followed by the opening of a water-filled pore that permits slow RNA diffusion, in line with the low efficiency and slow kinetics reported for endosomal escape. The rate-limiting step of endosomal escape arises from persistent electrostatic coupling between RNA and protonated ionizable lipids maintained by the immediate RNA-lipid environment, hindering cargo disengagement even after pore formation. This delayed release is consistent with the experimentally observed time-dependent inhibitory activity of the loaded LNP. Together, these results highlight the importance of local pK and protonation effects near the RNA, which are not captured by global apparent pK measurements.

biophysics↗

bioRxiv2024

Mechanism of expanded DNA recognition in xCas9

xCas9 is an evolved variant of the CRISPR-Cas9 genome editing system, engineered to improve specificity and reduce undesired off-target effects. How xCas9 expands the DNA targeting capability of Cas9 by recognizing a series of alternative Protospacer Adjacent Motif (PAM) sequences while ignoring others is unknown. Here, we elucidate the molecular mechanism underlying xCas9s expanded PAM recognition and provide critical insights for expanding DNA targeting. We demonstrate that while wild-type Cas9 enforces stringent guanine selection through the rigidity of its interacting arginine dyad, xCas9 introduces flexibility in R1335, enabling selective recognition of specific PAM sequences. This increased flexibility confers a pronounced entropic preference, which also improves recognition of the canonical TGG PAM. Furthermore, xCas9 enhances DNA binding to alternative PAM sequences during the early evolution cycles, while favouring binding to the canonical PAM in the final evolution cycle. This dual functionality highlights how xCas9 broadens PAM recognition and underscores the importance of fine-tuning the flexibility of the PAM-interacting cleft as a key strategy for expanding the DNA targeting potential of CRISPR-Cas systems. These findings deepen our understanding of DNA recognition in xCas9 and may apply to other CRISPR-Cas systems with similar PAM recognition requirements.

biophysics↗

bioRxiv2021

Determinants of directionality and efficiency of the ATP synthase Fo motor at atomic resolution

Fo subcomplex of ATP synthase is an membraneembedded rotary motor that converts proton motive force into mechanical energy. Despite a rapid increase in the number of high-resolution structures, the mechanism of tight coupling between proton transport and motion of the rotary c-ring remains elusive. Here, using extensive all-atom free energy simulations, we show how the motors directionality naturally arises from the interplay between intra-protein interactions and energetics of protonation of the c-ring. Notably, our calculations reveal that the strictly conserved arginine in the a-subunit (R176) serves as a jack-of-all-trades: it dictates the direction of rotation, controls the protonation state of the proton-release site and separates the two proton-access half-channels. Therefore, arginine is necessary to avoid slippage between the proton flux and the mechanical output and guarantees highly efficient energy conversion. We also provide mechanistic explanations for the reported defective mutations of R176, reconciling the structural information on the Fo motor with previous functional and single-molecule data.

biophysics↗