Differences in antigen‐binding affinity caused by single amino acid substitution in the variable region of the heavy chain

Immunology and Cell Biology - Tập 71 Số 4 - Trang 239-247 - 1993
Jim Xiang1, Zhi‐Nan Chen2, Louis T. J. Delbaere3, Enwu Liu1
1Saskatoon Cancer Center, Department of Microbiology and Division of Oncology, University of Saskatchewan, Saskatoon, Saskatchewan, Canada
2Department of Pathology, Fourth Medical University, Xian, China
3Department of Biochemistry, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.

Tóm tắt

SummarySite‐directed mutagenesis has been used to examine the importance of histidine‐99 in the VH CDR3 region of a mouse/human chimeric anti‐TAG72 antibody, cB72.3‐1‐3. The expression vectors mpSV2neo‐EPl‐Vm4–10C.γ1, containing seven different mutant VH region fragments (Vm4–10) in association with the immunoglobulin enhancer (E), promoter (P1) and human genomic Cγ1 region fragments, were transfected into a heavy chain loss mutant cell line B72.3Mut(K), respectively. Mutant chimeric antibodies cB72.3m4–10 were purified from the transfectant supernates, and their binding affinities for the TAG72 antigen relative to that of the original cB72.3‐1‐3 antibody were compared. Substitution of histidine‐99 by glutamine resulted in a higher affinity antibody (cB72.3m4) whereas substitution by isoleucine resulted in a lower affinity antibody (cB72.3m9). The binding affinity of these mutant antibodies varied nearly eight‐fold. It was concluded that the residue at position 99 in the VH CDR3 region is in a ‘contact’ position in the B72.3/TAG72 antibody‐combining site. The polar side‐chains of glutamine and asparagine or the ionized side‐chains of histidine, arginine or glutamic acid contribute to higher binding affinity, whereas the hydrophobic side‐chains of isoleucine, leucine or phenylalanine resulted in a lower binding affinity for the TAG72 antigen.

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Tài liệu tham khảo

10.1146/annurev.bi.48.070179.004525

Kabat E. A., 1987, Sequences of Proteins of Immunological Interest

10.1016/0022-2836(85)90137-8

10.1073/pnas.79.6.1979

10.1073/pnas.82.9.2945

10.1038/328731a0

10.1002/j.1460-2075.1988.tb03038.x

10.1073/pnas.87.12.4814

Thor A., 1986, Distribution of the oncofetal antigen, tumor–associated glycoprotein‐72 defined by the monoclonal antibody, B72.3, Cancer Res., 46, 3118

10.1016/0161-5890(90)90091-D

Xiang J., 1992, Genetic engineering of high affinity anti‐human colorectal tumor mouse/human chimeric antibody, Immunology, 75, 209

Kjeldsen T., 1988, Preparation and characterization of monoclonal antibodies directed to the tumor‐associated O‐linked sialosyl 2,6α‐N–acetyl‐galactosaminyl (Sialosyl‐Tn) epitope, Cancer Res., 48, 2214

10.1016/0076-6879(83)00074-9

10.1073/pnas.74.12.5463

Maniatis T., 1982, Molecular Cloning: A Laboratory Manual

10.1073/pnas.81.22.7161

Heyman B., 1984, An enzyme‐linked immunosorbent assay for measuring anti‐sheep erythrocyte antibodies, J Immunol. Methods, 68, 139, 10.1016/0022-1759(84)90150-9

10.1111/j.1749-6632.1949.tb27297.x

10.1002/j.1460-2075.1986.tb04522.x

10.1111/j.1600-065X.1987.tb00507.x

10.1073/pnas.83.8.2628

Sharon J., 1990, Structural characterization of idiotopes by using antibody variants generated by site‐directed mutagenesis, J. Immunol., 44, 4863, 10.4049/jimmunol.144.12.4863

Allen D., 1988, Modulation of antibody binding affinity by somatic mutation, Int. J. Cancer, 3, 1, 10.1002/ijc.2910410802

10.1126/science.2426778

10.1038/347483a0

10.1038/326358a0

10.1073/pnas.86.15.5938

10.1073/pnas.86.23.9268

10.1016/0161-5890(91)90097-4

Colcher D., 1988, Radio‐immunolocalization of human carcinoma xenografts with B72.3 second generation monoclonal antibodies, Cancer Res., 48, 4597

Molinolo A., 1990, Enhanced tumor binding using immunohistochemical analyses by second generation anti‐tumor‐associated glycoprotein 72 monoclonal antibodies versus monoclonal antibody B72.3 in human tissues, Cancer Res., 50, 1291

Schlom J., 1992, Therapeutic advantage of high‐affinity anticarcinoma radio‐immunoconjugates, Cancer Res., 52, 1067

Chapman P., 1990, Light chain variants of an IgG3 anti‐GD3 monoclonal antibody and the relationship among avidity, effector functions, tumor targeting, and antitumor activity, Cancer Res., 50, 1503

Springer G., 1983, Proposed molecular basis of murine tumor cell‐hepatocyte interaction, J. Biol. Chem., 258, 5702, 10.1016/S0021-9258(20)81949-8

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Immunology and Cell Biology

Tập 71 Số 4

239-247

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