Biochemical and immunological characterization of the DNA binding protein (RBP-J kappa) to mouse J kappa recombination signal sequence.
We have investigated whether J kappa recombination signal sequence (RS) binding protein (RBP-J kappa) has any partial catalytic activities involved in the VDJ recombination reaction, such as cleavage, ligation, and bending of DNA. Murine RBP-J kappa protein purified by J kappa-RS affinity chromatography did not show DNA cleavage activities but contained a strong DNA ligase activity. To obtain a large amount of purified RBP-J kappa protein, recombinant RBP-J kappa was synthesized in Escherichia coli as a fusion protein and also in silkworm cells. Although recombinant RBP-J kappa produced in silkworm cells could bind J kappa-RS, it failed to show either ligase or DNA bending activity. Since the DNA affinity-purified RBP-J kappa has the ligase activity, the RBP-J kappa protein may form a complex with a ligase in vivo. We have raised monoclonal antibodies against the RBP-J kappa fusion protein which was synthesized in E. coli and unable to bind J kappa-RS. Using the anti-RBP-J kappa monoclonal antibody we have shown that the RBP-J kappa protein is expressed ubiquitously in mammalian tissues. The ubiquitous expression of the RBP-J kappa protein is consistent with the hypothesis that the RBP-J kappa protein may have dual function [Furukawa et al. (1991) J. Biol. Chem. 266, 23334-23340].
Hamaguchi
Y
Y
Department of Medical Chemistry, Kyoto University Faculty of Medicine.
Yamamoto
Y
Y
Iwanari
H
H
Maruyama
S
S
Furukawa
T
T
Matsunami
N
N
Honjo
T
T
D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D002460:Cell Line; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007145:Immunoglobulin kappa-Chains / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D021382:Protein Sorting Signals / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011995:Recombination, Genetic
D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D007145:Immunoglobulin kappa-Chains; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D004247:DNA