CEM 2.0: Co-evolution-guided mining of superior extremophilic proteins via a curated proteome database and protein...

Extremophilic proteins offer immense potential for diverse applications; however, their discovery is hindered by the reliance on predefined templates and domain-specific knowledge. Therefore, new methods are required to mine extremophilic proteins without relying on existing templates, enabling a comprehensive exploration of functional extremophilic proteins across the entire protein universe. Here, we propose a co-evolution-guided protein mining framework, CEM 2.0 (Co-Evolution Mining 2.0), comprising three synergistic components: (1) an extensive and diverse extremophilic proteome database covering six extremophilic features, (2) a multidimensional evaluator for six extremophilic features, leveraging the protein language model ProteinBERT, and (3) an integrated and interactive mining pipeline equipped with advanced tools for computing >20 properties of proteins. This framework was implemented on an interactive web platform (https://cem.sjtu.edu.cn/) equipped with intuitive visualization tools. We conducted wet-lab validation, performed a series of computational validations using published studies, and made recommendations for novel PETases, thereby demonstrating the effectiveness and potential of CEM 2.0 for the template-independent discovery of extremophilic proteins. CEM 2.0 is expected to mine functional extremophilic proteins across the protein universe in terms of evolutionary distance and serve as a valuable tool for protein engineering and synthetic biology.