Dual domain recognition determines SARS-CoV-2 PLpro selectivity for human ISG15 and K48-linked di-ubiquitin.
Jerzy OsipiukPawel M WydorskiBenjamin T LanhamChristine TesarMichael EndresElizabeth EngleRobert JedrzejczakVishruth MullapudiKarolina MichalskaKrzysztof FidelisDavid FushmanAndrzej JoachimiakLukasz A JoachimiakPublished in: bioRxiv : the preprint server for biology (2022)
The Papain-like protease (PLpro) is a domain of a multi-functional, non-structural protein 3 of coronaviruses. PLpro cleaves viral polyproteins and posttranslational conjugates with poly-ubiquitin and protective ISG15, composed of two ubiquitin-like (UBL) domains. Across coronaviruses, PLpro showed divergent selectivity for recognition and cleavage of posttranslational conjugates despite sequence conservation. We show that SARS-CoV-2 PLpro binds human ISG15 and K48-linked di-ubiquitin (K48-Ub 2 ) with nanomolar affinity and detect alternate weaker-binding modes. Crystal structures of untethered PLpro complexes with ISG15 and K48-Ub 2 combined with solution NMR and cross-linking mass spectrometry revealed how the two domains of ISG15 or K48-Ub 2 are differently utilized in interactions with PLpro. Analysis of protein interface energetics uncovered differential binding stabilities of the two UBL/Ub domains. We emphasize how substrate recognition can be tuned to cleave specifically ISG15 or K48-Ub 2 modifications while retaining capacity to cleave mono-Ub conjugates. These results highlight alternative druggable surfaces that would inhibit PLpro function.
Keyphrases
- sars cov
- endothelial cells
- small molecule
- respiratory syndrome coronavirus
- mass spectrometry
- binding protein
- amino acid
- cancer therapy
- high resolution
- magnetic resonance
- protein protein
- pseudomonas aeruginosa
- cystic fibrosis
- escherichia coli
- capillary electrophoresis
- ms ms
- solid state
- gas chromatography
- candida albicans