Directed evolution of chemotaxis inhibitory protein of Staphylococcus aureus generates biologically functional variants with reduced interaction with human antibodies.
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.
Gustafsson
Erika
E
Alligator Bioscience AB, Scheelevägen 19A, S-223 70 Lund. erg@alligatorbioscience.com
Rosén
Anna
A
Barchan
Karin
K
van Kessel
Kok P M
KP
Haraldsson
Karin
K
Lindman
Stina
S
Forsberg
Cecilia
C
Ljung
Lill
L
Bryder
Karin
K
Walse
Björn
B
Haas
Pieter-Jan
PJ
van Strijp
Jos A G
JA
Furebring
Christina
C
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics* / Q000276:immunology* / Q000494:pharmacology; D002460:Cell Line; D019020:Directed Molecular Evolution; D015195:Drug Design; D015870:Gene Expression; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism; D044087:Receptor, Anaphylatoxin C5a / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics / Q000276:immunology / Q000494:pharmacology; D016415:Sequence Alignment; D013211:Staphylococcus aureus / Q000235:genetics / Q000276:immunology*
D001426:Bacterial Proteins; D007074:Immunoglobulin G; D044087:Receptor, Anaphylatoxin C5a; D011994:Recombinant Proteins; C493134:chemotaxis inhibitory protein, Staphylococcus aureus