Affinity maturation of human CD4 by yeast surface display and crystal structure of a CD4HLA-DR1 complex
Lili Wang, Xin Xiang Wang, Y Li, Min Mo, Yiyuan Yi, Qian Wang, W Gao, Roy A. Mariuzza
Helper T cell activation generally requires the co-receptor CD4, which binds MHC class II molecules. A remarkable feature of the CD4MHC class II interaction is its exceptionally low affinity, which ranges from KD ~200 μM to >2 mM. Investigating the biological role of the much lower affinity of this interaction than those of other cellcell recognition molecules will require CD4 mutants with enhanced binding to MHC class II for testing in models of T cell development. To this end, we used in vitro directed evolution to increase the affinity of human CD4 for HLADR1. A mutant CD4 library was displayed on the surface of yeast and selected using HLA-DR1 tetramers or monomers, resulting in isolation of a CD4 clone containing 11 mutations. Reversion mutagenesis showed that most of the affinity increase derived from just two substitutions, Gln40Tyr and Thr45Trp. A CD4 variant bearing these mutations bound HLA-DR1 with KD = 8.8 μM, compared to >400 μM for wild-type CD4. To understand the basis for improved affinity, we determined the structure of this CD4 variant in complex with HLA-DR1 to 2.4 Å resolution. The structure provides the first atomic-level description of the CD4-binding site on MHC class II and reveals how CD4 recognizes highly polymorphic HLA-DR, -DP, and -DQ molecules by targeting invariant residues in their α2 and β2 domains. Additionally, the CD4 mutants reported here constitute unique tools for probing the influence of CD4 affinity on T cell activation and development.
Proceedings of the National Academy of Sciences of the United States of America
, , X.
, Li, Y.
, Mo, M.
, Yi, Y.
, Wang, Q.
, Gao, W.
and Mariuzza, R.
Affinity maturation of human CD4 by yeast surface display and crystal structure of a CD4–HLA-DR1 complex, Proceedings of the National Academy of Sciences of the United States of America, [online], https://tsapps.nist.gov/publication/get_pdf.cfm?pub_id=908864
(Accessed December 9, 2023)