Biology subjects

Mancino, A.

Publications and source records attributed to Mancino, A..

3 recordsLinked to original sources

bioRxiv2026

Cryo-EM of a nucleotide-polymerizing ribozyme enables its predictive improvement

Ribozymes capable of self-replication from nucleotides would have been central to the hypothesized RNA World. The leading laboratory models for such molecules were converted from a class I ligase by in vitro evolution but then developed without 3D structures. Here, scaffolded cryo-EM of the substrate-free tC19Z RNA polymerase ribozyme at 3.1 [A] resolution shows how this conversion was achieved. An accessory domain evolved from random sequence grips the ancestral ligase through a loop-loop contact, a seam of magnesium ions, and a six-base stack, and rebuilds the ligases substrate binding site from different residues of its own. A previously unrecognized pairing, present before substrate binds, sequesters the 5' end that must otherwise pair with the template. Compensatory mutations to the ribozyme and template, designed to break this ectopic pairing, increase the extension rate. These results suggest that accelerating RNA structure determination may speed progress toward nucleotide-based self-replication.

biochemistry↗

bioRxiv2026

De novo design of RNA pseudoknots with deep learning

RNA design has been hindered by the limited accuracy of 3D structure prediction. Here, we show that intricate RNA structures can be generated with current deep learning tools through accurate de novo design of pseudoknot secondary structures. In an Eterna competition involving 57 pseudoknots, generative AI methods matched experienced human designers in solving most blind challenges, evaluated by single-nucleotide-resolution chemical mapping, compensatory mutagenesis, and cryogenic electron microscopy. AI-generated molecules with accurate secondary structures formed well-ordered 3D folds stabilized by noncanonical tertiary interactions not modeled during design. Success was guided by an RNet foundation model trained on prior chemical mapping data, suggesting that some difficult RNA design tasks may be tractable without first solving RNA 3D structure prediction.

biophysics↗

bioRxiv2021

CryoEM and AI reveal a structure of SARS-CoV-2 Nsp2, a multifunctional protein involved in key host processes.

The SARS-CoV-2 protein Nsp2 has been implicated in a wide range of viral processes, but its exact functions, and the structural basis of those functions, remain unknown. Here, we report an atomic model for full-length Nsp2 obtained by combining cryo-electron microscopy with deep learning-based structure prediction from AlphaFold2. The resulting structure reveals a highly-conserved zinc ion-binding site, suggesting a role for Nsp2 in RNA binding. Mapping emerging mutations from variants of SARS-CoV-2 on the resulting structure shows potential host-Nsp2 interaction regions. Using structural analysis together with affinity tagged purification mass spectrometry experiments, we identify Nsp2 mutants that are unable to interact with the actin-nucleation-promoting WASH protein complex or with GIGYF2, an inhibitor of translation initiation and modulator of ribosome-associated quality control. Our work suggests a potential role of Nsp2 in linking viral transcription within the viral replication-transcription complexes (RTC) to the translation initiation of the viral message. Collectively, the structure reported here, combined with mutant interaction mapping, provides a foundation for functional studies of this evolutionary conserved coronavirus protein and may assist future drug design.

biophysics↗