Structure-based design of an rnase chimera for antimicrobial therapy

Guillem Prats-Ejarque*, Helena Lorente, Clara Villalba, Raúl Anguita, Lu Lu, Sergi Vázquez-Monteagudo, Pablo Fernández-Millán, Ester Boix

*Corresponding author for this work

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Bacterial resistance to antibiotics urges the development of alternative therapies. Based on the structure-function of antimicrobial members of the RNase A superfamily, we have developed a hybrid enzyme. Within this family, RNase 1 exhibits the highest catalytic activity and the lowest cytotoxicity; in contrast, RNase 3 shows the highest bactericidal action, alas with a reduced catalytic activity. Starting from both parental proteins, we designed a first RNase 3/1-v1 chimera. The construct had a catalytic activity much higher than RNase 3, unfortunately without reaching an equivalent antimicrobial activity. Thus, two new versions were created with improved antimicrobial properties. Both of these versions (RNase 3/1-v2 and-v3) incorporated an antimicrobial loop characteristic of RNase 3, while a flexible RNase 1-specific loop was removed in the latest construct. RNase 3/1-v3 acquired both higher antimicrobial and catalytic activities than previous versions, while retaining the structural determinants for interaction with the RNase inhibitor and displaying non-significant cytotoxicity. Following, we tested the constructs’ ability to eradicate macrophage intracellular infection and observed an enhanced ability in both RNase 3/1-v2 and v3. Interestingly, the inhibition of intracellular infection correlates with the variants’ capacity to induce autophagy. We propose RNase 3/1-v3 chimera as a promising lead for applied therapeutics.

Original languageEnglish
Article number95
Number of pages21
JournalInternational journal of molecular sciences
Volume23
Issue number1
DOIs
Publication statusPublished - 1 Jan 2022

Keywords

  • Antimicrobial proteins
  • Protein engineering
  • RNase
  • Structure-function relationship

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