Effects of substitutions of closely related amino acids at the contact surface in an antigen-antibody complex on thermodynamic parameters.
We constructed a library of 512 kinds of Fv fragment, derivatives of a monoclonal antibody, D1.3, specific for hen egg-white lysozyme, in which a total of nine of the original amino acids were replaced by closely related amino acids at positions in the complementarity-determining regions of the H chain. More than 80% of the clones in the library produced Fv fragments in Escherichia coli. Two wild-type and 13 mutant Fv fragments were prepared in large quantities and subjected to analysis by differential titration calorimetry. The association constants of the 15 Fv fragments with hen egg-white lysozyme were distributed between 0.12 x 10(7) and 1.59 x 10(8) M-1. The changes in delta H0 and -T delta S0 caused by one-point mutation at each position did not have intrinsic values for each change. The same changes at one position had different effects on KA, delta H0, and -T delta S0 when differences had been introduced in other regions. The delta(delta G0) caused by a single-point mutation ranged from -0.56 to 1.56 kcal/mol. By contrast, the delta(delta H0) and delta(-T delta S0) caused by a single-point mutation ranged from -3.5 to 3.4 and from -3.8 to 3.4 kcal/mol, respectively. When antibodies gain the binding energy contributed by the effects of enthalpy, they lose the binding energy contributed by the effects of entropy and vice versa. In general, changes in entropy compensate for changes in enthalpy.
Ito
W
W
Institute for Comprehensive Medical Science, Fujita Health University, Aichi, Japan.
Iba
Y
Y
Kurosawa
Y
Y
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D000936:Antigen-Antibody Complex / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D004247:DNA; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D008969:Molecular Sequence Data; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D013816:Thermodynamics
D000596:Amino Acids; D000936:Antigen-Antibody Complex; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D004247:DNA