Exploring the RNase A scaffold to combine catalytic and antimicrobial activities. Structural characterization of RNase 3/1 chimeras
Fernández-Millán, Pablo (Universitat Autònoma de Barcelona. Departament de Bioquímica i de Biologia Molecular)
Boix i Borràs, Esther (Universitat Autònoma de Barcelona. Departament de Bioquímica i de Biologia Molecular)
Prats-Ejarque, Guillem (Universitat Autònoma de Barcelona. Departament de Bioquímica i de Biologia Molecular)
Vázquez-Monteagudo, Sergi (Universitat Autònoma de Barcelona. Departament de Bioquímica i de Biologia Molecular)

Data:	2022
Resum:	Design of novel antibiotics to fight antimicrobial resistance is one of the first global health priorities. Novel protein-based strategies come out as alternative therapies. Based on the structure-function knowledge of the RNase A superfamily we have engineered a chimera that combines RNase 1 highest catalytic activity with RNase 3 unique antipathogen properties. A first construct (RNase 3/1-v1) was successfully designed with a catalytic activity 40-fold higher than RNase 3, but alas in detriment of its anti-pathogenic activity. Next, two new versions of the original chimeric protein were created showing improvement in the antimicrobial activity. Both second generation versions (RNases 3/1-v2 and -v3) incorporated a loop characteristic of RNase 3 (L7), associated to antimicrobial activity. Last, removal of an RNase 1 flexible loop (L1) in the third version enhanced its antimicrobial properties and catalytic efficiency. Here we solved the 3D structures of the three chimeras at atomic resolution by X-ray crystallography. Structural analysis outlined the key functional regions. Prediction by molecular docking of the protein chimera in complex with dinucleotides highlighted the contribution of the C-terminal region to shape the substrate binding cavity and determine the base selectivity and catalytic efficiency. Nonetheless, the structures that incorporated the key features related to RNase 3 antimicrobial activity retained the overall RNase 1 active site conformation together with the essential structural elements for binding to the human ribonuclease inhibitor (RNHI), ensuring non-cytotoxicity. Results will guide us in the design of the best RNase pharmacophore for anti-infective therapies.
Ajuts:	Ministerio de Economía y Competitividad SAF 2015-66007P Agencia Estatal de Investigación PID 2019-106123GB-I00 Fundació la Marató de TV3 2080310 Agència de Gestió d'Ajuts Universitaris i de Recerca 2019/LLAV-00002
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	RNase 3 ; RNase 1 ; Chimera ; Crystal structure ; Catalytic activity ; Base binding sites ; Antimicrobial protein
Publicat a:	Frontiers in Molecular Biosciences, Vol. 9 (September 2022) , art. 964717, ISSN 2296-889X

DOI: 10.3389/fmolb.2022.964717
PMID: 36188223

17 p, 4.4 MB

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