Metal binding properties of a monoclonal antibody directed toward metal-chelate complexes.
A monoclonal antibody that recognizes cadmium-EDTA complexes has been produced by the injection of BALB/c mice with a metal-chelate complex covalently coupled to a carrier protein. The ability of purified antibody to recognize 16 different metal-EDTA complexes was assessed by measuring equilibrium binding constants using a KinExATM immunoassay instrument. The antibody bound to cadmium- and mercury-EDTA complexes with equilibrium dissociation constants of 21 and 26 nM, respectively. All other metal-EDTA complexes tested, including those of Mn(II), In(III), Ni(II), Zn(II), Co(II), Cu(II), Ag(I), Fe(III), Pb(II), Au(III), Tb(III), Ga(III), Mg(II), and Al(III) bound with affinities from 20- to 40,000-fold less than that determined for the cadmium-EDTA complex. With the exception of mercury and magnesium, the binding of divalent metal-chelate complexes was well-correlated with the size of the metal ion. The amino acid sequences of the heavy and light chain variable regions were deduced from polymerase chain reaction-amplified regions of the corresponding genes and subsequently used to construct molecular models of the antigen binding region. The key residue for cadmium binding in the model for 2A81G5 appeared to be histidine 96 in the heavy chain.
Department of Ophthalmology, Tulane University School of Medicine, New Orleans, Louisiana 70112, USA. dblake@tmcpop.tmc.tulane.edu
D A
DA
Blake
P
P
Chakrabarti
M
M
Khosraviani
F M
FM
Hatcher
C M
CM
Westhoff
P
P
Goebel
D E
DE
Wylie
R C
RC
Blake
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics; D000915:Antibody Affinity; D001483:Base Sequence; D002412:Cations / Q000276:immunology; D002413:Cations, Divalent / Q000276:immunology; D003001:Cloning, Molecular; D004492:Edetic Acid; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry; D007147:Immunoglobulin Light Chains / Q000737:chemistry / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics; D008670:Metals / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D000911:Antibodies, Monoclonal; D002412:Cations; D002413:Cations, Divalent; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D008670:Metals; D011994:Recombinant Proteins; D004492:Edetic Acid