Biology subjects

Fatehi, F.

Publications and source records attributed to Fatehi, F..

3 recordsLinked to original sources

bioRxiv2024

Point mutation in a virus-like capsid drives symmetry reduction to form tetrahedral cages

Protein capsids are a widespread form of compartmentalisation in nature. Icosahedral symmetry is ubiquitous in capsids derived from spherical viruses, as this geometry maximises the internal volume that can be enclosed within. Despite the strong preference for icosahedral symmetry, we show that simple point mutations in a virus-like capsid can drive the assembly of novel symmetry-reduced structures. Starting with the encapsulin from Myxococcus xanthus, a 180-mer bacterial capsid that adopts the well-studied viral HK97 fold, we use mass photometry and native charge detection mass spectrometry to identify a triple histidine point mutant that forms smaller dimorphic assemblies. Using cryo-EM, we determine the structures of a precedented 60-mer icosahedral assembly and an unprecedented 36-mer tetrahedron that features significant geometric rearrangements around a novel interaction surface between capsid protomers. We subsequently find that the tetrahedral assembly can be generated by triple point mutation to various amino acids, and that even a single histidine point mutation is sufficient to form tetrahedra. These findings represent the first example of tetrahedral geometry across all characterised encapsulins, HK97-like capsids, or indeed any virus-derived capsids reported in the Protein Data Bank, revealing the surprising plasticity of capsid self-assembly that can be accessed through minimal changes in protein sequence. Significance statementViral capsids are cage-like protein assemblies that preferentially adopt icosahedral symmetry to maximise their internal volume for housing genetic material. This icosahedral preference extends to encapsulins, a widespread family of bacterial protein cages which evolved from viral capsids. Counter to this fundamental geometric preference, the formation of well-defined tetrahedral cages from a single amino acid substitution in an encapsulin reveals the surprising geometric flexibility of a common viral protein fold. These findings suggest that protein oligomerisation is far more permissive than intuitively expected, where serendipitous interactions between proteins arising from minimal mutations can cascade to form vast architectural changes. The ability to redesign protein architectures through simple mutations should enable biotechnological advances in vaccine development, drug delivery, and enzymatic biomanufacturing.

biochemistry↗

bioRxiv2023

Local structural flexibility drives oligomorphism in computationally designed protein assemblies

Many naturally occurring protein assemblies have dynamic structures that allow them to perform specialized functions. For example, clathrin coats adopt a wide variety of architectures to adapt to vesicular cargos of various sizes. Although computational methods for designing novel self-assembling proteins have advanced substantially over the past decade, most existing methods focus on designing static structures with high accuracy. Here we characterize the structures of three distinct computationally designed protein assemblies that each form multiple unanticipated architectures, and identify flexibility in specific regions of the subunits of each assembly as the source of structural diversity. Cryo-EM single-particle reconstructions and native mass spectrometry showed that only two distinct architectures were observed in two of the three cases, while we obtained six cryo-EM reconstructions that likely represent a subset of the architectures present in solution in the third case. Structural modeling and molecular dynamics simulations indicated that the surprising observation of a defined range of architectures, instead of non-specific aggregation, can be explained by constrained flexibility within the building blocks. Our results suggest that deliberate use of structural flexibility as a design principle will allow exploration of previously inaccessible structural and functional space in designed protein assemblies.

bioengineering↗

bioRxiv2021

Dysregulation of Hepatitis B Virus Nucleocapsid Assembly with RNA-directed Small Ligands

RNA sequences/motifs dispersed across the pre-genomic copy of the Hepatitis B Virus genome regulate formation of nucleocapsids in vitro in an epsilon/polymerase independent fashion. These multiple RNA Packaging Signals (PSs) form stem-loops presenting in each loop a core protein recognition motif, -RGAG-. Small, drug-like molecules binding these motifs were identified by screening an immobilized library with a fluorescently-labelled RNA oligonucleotide encompassing the most conserved of these sites. This identified 66 "hits", with affinities ranging from low nanomolar to high micromolar in SPR assays. High affinity ligand binding is dependent on the presence of the -RGAG-motif, which also appears to be the common element in cross-binding to other PS sites. Some of these compounds are potent inhibitors of in vitro core protein assembly around the HBV pre-genome. Mathematical modelling confirms the potential of these novel anti-viral drug targets for disrupting replication of this major human pathogen. Preliminary structure-activity relationships of the highest affinity compound reveal critical functional groups for PS-binding. PS-regulated assembly is easily adapted to high-throughput screening allowing future development of pharmacologically active compounds.

microbiology↗