An isotype switched and somatically mutated rheumatoid factor clone isolated from a MRL-lpr/lpr mouse exhibits limited intraclonal affinity maturation.
Employing site-directed mutagenesis we have reconstructed and expressed the germ-line precursor of an expanded rheumatoid factor (RF) clone. This RF clone, designated clone F, was isolated from an autoimmune MRL/MpJ-lpr/lpr mouse. Most of the clone members were extensively mutated and isotyped-switched. The predominant isotype of clone F was gamma 3. The RF bound specifically to the MRL gamma 2 a allotype (Igh-1j) but not to the B6 gamma 2a allotype (Igh-1b). The germ-line antibody was also found to bind gamma 2a in an RF assay. The affinities of the germ-line RF and representative members of the clone were measured in an ELISA-based equilibrium binding assay. The dissociation constant (Kd) of the germ-line RF was 2.5 x 10(-6) M. All of the expressed clone members had affinities within a two- to sixfold range of the germ line, indicating that the mechanisms of somatic hypermutation and selection resulted in only limited affinity maturation of this autoantibody clone.
Jacobson
B A
BA
Department of Pathology, Boston University School of Medicine, MA 02118.
Sharon
J
J
Shan
H
H
Shlomchik
M
M
Weigert
M G
MG
Marshak-Rothstein
A
A
D000595:Amino Acid Sequence; D000818:Animals; D000915:Antibody Affinity; D000916:Antibody Diversity / Q000235:genetics; D001402:B-Lymphocytes / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D006825:Hybridomas / Q000276:immunology; D017578:Immunoglobulin Class Switching / Q000235:genetics*; D007700:Kinetics; D051379:Mice; D008817:Mice, Mutant Strains; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D012217:Rheumatoid Factor / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D014162:Transfection
D018076:DNA, Complementary; D012217:Rheumatoid Factor