https://scholars.lib.ntu.edu.tw/handle/123456789/202191
Title: | Discontinuous Residues of Factor Ix Constitute a Surface for Binding the Anti-Factor Ix Monoclonal Antibody a-5 | Authors: | Chang, Yu-Jia Wu, Hua-Lin Hsu, Ya-Chu Hamaguchi, Nobuko Shi, Guey-Yueh Shen, Ming-Ching Lin, Shu-Wha |
Keywords: | factor IX;alanine-scanning mutagenesis;recombinant proteins;monoclonal antibody;conformational epitope;surface plasmon resonance | Issue Date: | 2003 | Journal Volume: | v.111 | Journal Issue: | n.4-5 | Start page/Pages: | 293-299 | Source: | THROMBOSIS RESEARCH | Abstract: | Anti-human factor IX monoclonal antibody, A-5 (Mab A-5), has been widely used in structure-function studies for factor IX. Mab A-5 recognizes the catalytic domain of human factor IX (FIX). Regions important for Mab A-5 binding have previously been localized to the amino terminus of the heavy chain of factor IX, encompassing amino acid residues 181- 310 [Blood (74) 971]. We have selected 20 positions in this region for alanine-scanning mutagenesis. We found that Mab A -5 failed to react with the recombinant factor IX mutants with substitutions at positions 228 and 252. Mab A-5 also reacted to a lesser extent to FIXD276A (factor IX with alanine substitution for aspartic acid at residue 276) and FIXK201A/D203A (double alanine substitutions at residues 201 and 203). The apparent dissociation rate constants (K-D) in binding Mab A-5 were 6.0 x 10(-9), 1.4 x 10(-8) and 2.0 x 10(-8) M, for wild-type FIX, FIXK201A/D203A and FIXD276A, respectively. The increased K-D values of the two FIX mutants are mainly owing to the increased dissociation rates . These affected residues constitute a surface opposite from the factor VIIIa binding surface. We conclude that the epitope for Mab A-5 is on a surface composed of residues 228 , 252, 276, and 201 or 203. This surface, which may not be important for factor VIII binding, contains a strong antigenic region on factor IX. (C) 2003 Elsevier Ltd. All rights reserved |
URI: | http://ntur.lib.ntu.edu.tw//handle/246246/103381 |
Appears in Collections: | 醫學系 |
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