This research is led by Prof. Wen-Bin Zhang (Faculty of Chemistry and Molecular Engineering, Peking College & Beijing Academy of Synthetic Intelligence) and Dr. Jing Fang (Faculty of Chemistry and Molecular Engineering, Peking College). A single-domain protein catenane refers to 2 mechanically interlocked polypeptide rings that fold synergistically right into a compact and built-in construction, which is extraordinarily uncommon in nature. This design was achieved by rewiring the connectivity between secondary motifs to introduce synthetic entanglement, and synthesis was readily completed via a sequence of programmed streamlined post-translational processing occasions in cells with none extra in vitro reactions.
The one-domain catenane cat-DHFR was totally characterised. Proof from mixed SDS-PAGE, SEC, LC-MS, IMS-MS, and proteolytic digestion experiments unambiguously proved its topology. The cat-DHFR displays enhanced anti-aggregation properties and has a Tm that’s 6 °C increased than the linear management. Though the catalytic exercise of cat-DHFR is diminished owing to its decreased affinity towards the substrate and cofactor, it has higher thermal resilience than l-DHFR. Even after incubation at 70 °C for 10 min, cat-DHFR retained over 70% of the catalytic exercise, whereas the linear management misplaced virtually all exercise. The analysis crew anticipate that this methodology might be typically relevant to different single-domain proteins, together with these with folds much like DHFR or with fully totally different folds. The provision of those single-domain protein catenanes facilitates the elucidation of topological results on construction–property relationships. The outcomes additional suggest that it’s attainable to map the present linear protein universe into single-domain protein catenanes with well-preserved features and extra advantages, opening up new territory for protein molecules. Surpassing the linear paradigm of pure protein molecules, these topological proteins are multi-chain, multi-dimensional molecules with purposeful advantages of topology, wealthy design risk and wonderful evolvability. As a brand new class of protein molecules, they maintain nice potential for a broad vary of purposes, together with, however not restricted to, industrial enzymes, antibodies, cytokines, and biomaterials.
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Journal reference:
Fang, J., et al. (2023). A Single-domain Protein Catenane of Dihydrofolate Reductase. Nationwide Science Overview. doi.org/10.1093/nsr/nwad304.
