X‐ray structures of fragments from binding and nonbinding versions of a humanized anti‐CD18 antibody: Structural indications of the key role of VH residues 59 to 65

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Abstract
X‐ray crystal structures of fragments from two different humanized antiCD18 antibodies are reported. The Fv fragment of the nonbinding version has been refined in space group C2 with a=64.2 Å, b=61.3 Å, c=51.8 Å, and β=99° to an R‐value of 18.0% at 1.9 Å, and the Fab fragment of the tight‐binding version has been refined in space group P3 with a=101. Å and c=45.5 Å to an R‐value of 17.8% at 3.0 Å resolution. The very large difference in their binding affinity (>1000‐fold) is attributed to large and local structural differences in the C‐terminal part of CDR‐H2, and from this we conclude there is direct contact between this region and antigen when they combine. X‐ray structures of antibody–antigen complexes available in the literature have yet to show this part of CDR‐H2 in contact with antigen, despite its hypervariable sequence. Implications of this result for antibody humanization are discussed. © 1994 John Wiley & Sons, Inc.