Biology subjects

Colizzi, F.

Publications and source records attributed to Colizzi, F..

3 recordsLinked to original sources

bioRxiv2026

A cold-adapted Cas9 nuclease from the deep ocean

The abundance of CRISPR systems in bacteria and archaea fuels the search for Cas nucleases that perform where established enzymes fail. Deep Ocean 1 Cas9 (DO1Cas9) is to our knowledge the first Cas9 nuclease characterized from the deep ocean, recovered from a metagenomic sample at 1,500 m depth in the Atlantic Ocean. DO1Cas9 shows cold-adapted structural flexibility, thermal susceptibility, salt-dependent stability, higher specificity than SpCas9 and cleaves double-stranded DNA efficiently under cold conditions where mesophilic SpCas9 retains only residual activity. DO1Cas9 edits genomes in diverse organisms such as Caenorhabditis elegans, zebrafish, the marine tunicate Oikopleura dioica, and the malaria parasite Plasmodium falciparum. In the marine bacterium Marinomonas mediterranea, DO1Cas9 programmed against an invading phage reduced plaquing efficiency about 10-fold more than SpCas9. These results expand the CRISPR-Cas9 toolbox with a nuclease for cold-chain editing and marine biotechnology.

genetics↗

bioRxiv2025

Energetic Constraints in the Enzymatic Depolymerization of Crystalline PET

Polyethylene terephthalate (PET) is a widely used thermoplastic whose high crystallinity poses a major barrier to upscaling enzymatic recycling. While PETases with high activity and stability have been reported, no enzyme capable of directly depolymerizing crystalline PET (cPET) has been discovered, and the molecular determinants limiting their efficacy remain difficult to characterize. Here, we integrate experimental conformational ratios of crystalline and amorphous PET chains with enhanced-sampling molecular dynamics simulations to map the free-energy landscape of a prototypical PETase bound to PET oligomers, revealing how structural equilibria translate to catalytic function. Surprisingly, productive enzyme-substrate catalytic configurations can be reached for both crystalline and amorphous PET chains. However, forming catalytic ensembles with cPET requires ~25 kJ/mol more than with aPET, with an additional ~17 kJ/mol per monomer needed for chain separation, which further limits enzymatic activity on crystalline substrates. The model highlights limitations of current alpha/beta-hydrolase scaffolds used for PET depolymerization and indicates directions for their redesign to enable cPET depolymerization. Our approach showcases a general strategy to explore substrate-enzyme catalytic ensembles in plastic depolymerization and guide enzyme design with built-in sustainability.

biophysics↗

bioRxiv2022

Prediction Of The Impact Of Genetic Variability On Drug Sensitivity For Clinically Relevant EGFR Mutations

Mutations in the kinase domain of the Epidermal Growth Factor Receptor (EGFR) can be drivers of cancer and also trigger drug resistance in patients under chemotherapy treatment based on kinase inhibitors use. A priori knowledge of the impact of EGFR variants on drug sensitivity would help to optimize chemotherapy and to design new drugs effective against resistant variants. To this end, we have explored a variety of in silico methods, from sequence-based to state-of-the-art atomistic simulations. We did not find any sequence signal that can provide clues on when a drug-related mutation appears and what will be the impact in drug activity. Low-level simulation methods provide limited qualitative information on regions where mutations are likely to produce alterations in drug activity and can predict around 70% of the impact of mutations on drug efficiency. High-level simulations based on non-equilibrium alchemical free energy calculations show predictive power. The integration of these state-of-the-art methods in a workflow implementing an interface for parallel distribution of the calculations allows its automatic and high-throughput use, even for researchers with moderate experience in molecular simulations.

biochemistry↗