Biology subjects

Elena-Real, C. A.

Publications and source records attributed to Elena-Real, C. A..

4 recordsLinked to original sources

bioRxiv2026

Emergence of new function through evolutionary divergence of an intrinsically disordered region

Intrinsically disordered regions (IDRs) are major drivers of protein functional diversification, yet the molecular features that enable the emergence of new functions within disordered sequences remain poorly understood. FCHO1 and FCHO2 are paralogous pioneer proteins of clathrin-mediated endocytosis that share a conserved domain architecture but perform distinct cellular functions and cannot compensate for each other's loss. Here we show that functional divergence between these proteins is associated with the acquisition of transient structure within their disordered regions. Using nuclear magnetic resonance spectroscopy, we identify two highly populated -helical elements in the IDR of FCHO1 that are absent from FCHO2. One of these helices mediates FCHO1 self-association and drives intracellular assembly, whereas FCHO2 lacks this behavior. Introduction of the FCHO1 helix into FCHO2 is sufficient to confer self-association and cellular assembly, demonstrating that a transient structural element embedded within an IDR can act as a transferable functional module. Evolutionary analysis reveals that this helical propensity emerged following duplication of the ancestral FCHO gene and became progressively reinforced during evolution. Despite this divergence, the same region retains a conserved membrane-binding activity in both paralogs. Together, our findings show how acquisition of transient secondary structure within an intrinsically disordered region can generate new molecular behaviors while preserving ancestral functions, providing a mechanism for the functional specialization of paralogous proteins.

biophysics↗

bioRxiv2023

Site-Specific Introduction of Alanines for the NMR Investigation of Low-Complexity Regions and Large Biomolecular Assemblies

NMR studies of large biomolecular machines and highly repetitive proteins remain challenging due to the difficulty of assigning signals to individual nuclei. Here, we present an efficient strategy to address this challenge by engineering a Pyrococcus horikoshii tRNA/alanyl-tRNA synthetase pair that enables the incorporation of up to three isotopically labeled alanine residues in a site-specific manner using in vitro protein expression. We have demonstrated the general applicability of this approach for NMR assignment by introducing isotopically labeled alanines into four proteins, including the 300-kDa molecular chaperone ClpP and the alanine-rich Phox2B transcription factor. For large protein assemblies, our labeling approach enables unambiguous assignments, while avoiding potential artefacts induced by site-specific mutations. When applied to Phox2B, which contains two poly-alanine tracts of nine and twenty alanines, we observe that the helical stability is strongly dependent on the homorepeat length, demonstrating structural cooperativity. The capacity to selectively introduce alanines with distinct labeling patterns is a powerful tool to probe structure and dynamics of biomolecular systems that are out of the reach of traditional structural biology methods.

biochemistry↗

bioRxiv2022

The structure of pathogenic huntingtin exon-1 defines the bases of its aggregation propensity

Huntingtons Disease is a neurodegenerative disorder caused by a CAG expansion of the first exon of the HTT gene, resulting in an extended poly-glutamine (poly-Q) tract in the N-terminus of the protein huntingtin (httex1). The structural changes occurring to the poly-Q when increasing its length remain poorly understood mainly due to its intrinsic flexibility and the strong compositional bias of the protein. The systematic application of site-specific isotopic labeling has enabled residue-specific NMR investigations of the poly-Q tract of pathogenic httex1 variants with 46 and 66 consecutive glutamines. The integrative analysis of the data reveals that the poly-Q tract adopts long -helical conformations stabilized by glutamine side-chain to backbone hydrogen bonds. 19F-NMR of site-specifically incorporated fluoro-glutamines and molecular dynamics simulations demonstrate that the mechanism propagating -helical conformations towards the poly-Q from the upstream N17 domain is independent of the poly-Q track length. Aggregation and atomic force microscopy experiments show that the presence of long and persistent -helices in the poly-Q tract is a stronger signature in defining the aggregation kinetics and the structure of the resulting fibrils than the number of glutamines. The ensemble of our observations provides a structural perspective of the pathogenicity of expanded httex1 and paves the way to a deeper understanding of poly-Q related diseases.

biophysics↗

bioRxiv2020

Mitochondrial cytochrome c liberates the nucleophosmin-sequestered ARF tumor suppressor in the nucleolus

The alternative reading frame (ARF) protein is crucial in the cellular response to oncogenic stress, being likewise the second most frequently inactivated gene in a wide spectrum of human cancers. ARF is usually sequestered in the nucleolus by the well-known oncogenic nucleophosmin (NPM) protein and is liberated in response to cell damage to exhibit its tumor-suppressor ability. However, the mechanism underlying ARF activation is unknown. Here we show that mitochondria-to-nucleus translocation of cytochrome c upon DNA damage leads to the break-off of the NPM-ARF ensemble and subsequent release of ARF from the nucleoli. Our structural and subcellular data support a molecular model in which the hemeprotein triggers the extended-to-compact conformation of NPM and competes with ARF for binding to NPM.

biophysics↗