Directed evolution of chemotaxis inhibitory protein of Staphylococcus aureus generates biologically functional variants with reduced interaction with human antibodies

Protein Eng Des Sel. 2010 Feb;23(2):91-101. doi: 10.1093/protein/gzp062. Epub 2009 Dec 3.

Abstract

Chemotaxis inhibitory protein of Staphylococcus aureus (CHIPS) is a protein that binds and blocks the C5a receptor (C5aR) and formylated peptide receptor, thereby inhibiting the immune cell recruitment associated with inflammation. If CHIPS was less reactive with existing human antibodies, it would be a promising anti-inflammatory drug candidate. Therefore, we applied directed evolution and computational/rational design to the CHIPS gene in order to generate new CHIPS variants displaying lower interaction with human IgG, yet retaining biological function. The optimization was performed in four rounds: one round of random mutagenesis to add diversity into the CHIPS gene and three rounds of DNA recombination by Fragment INduced Diversity (FIND). Every round was screened by phage selection and/or ELISA for decreased interaction with human IgG and retained C5aR binding. The mean binding of human anti-CHIPS IgG decreased with every round of evolution. For further optimization, new amino acid substitutions were introduced by rational design, based on the mutations identified during directed evolution. Finally, seven CHIPS variants with low interaction with human IgG and retained C5aR blocking capacity could be identified.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / analysis
  • Bacterial Proteins / genetics*
  • Bacterial Proteins / immunology*
  • Bacterial Proteins / pharmacology
  • Cell Line
  • Directed Molecular Evolution*
  • Drug Design
  • Gene Expression
  • Humans
  • Immunoglobulin G / immunology*
  • Models, Molecular
  • Molecular Sequence Data
  • Neutrophils / drug effects
  • Neutrophils / metabolism
  • Receptor, Anaphylatoxin C5a / antagonists & inhibitors*
  • Receptor, Anaphylatoxin C5a / metabolism
  • Recombinant Proteins / analysis
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Recombinant Proteins / pharmacology
  • Sequence Alignment
  • Staphylococcus aureus / genetics
  • Staphylococcus aureus / immunology*

Substances

  • Bacterial Proteins
  • Immunoglobulin G
  • Receptor, Anaphylatoxin C5a
  • Recombinant Proteins
  • chemotaxis inhibitory protein, Staphylococcus aureus