糖基化
分泌物
重组DNA
突变体
化学
免疫球蛋白E
生物活性
生物化学
生物
抗体
体外
免疫学
基因
作者
Robert J. Young,Raymond J. Owens,Graham A. Mackay,Christina M.W. Chan,Jimin Shi,Michihiro Hide,David M. Francis,Alistair J. Henry,Brian J. Sutton,Hannah J. Gould
出处
期刊:Protein Engineering Design & Selection
[Oxford University Press]
日期:1995-01-01
卷期号:8 (2): 193-199
被引量:50
标识
DOI:10.1093/protein/8.2.193
摘要
We have constructed an expression vector that leads to secretion of the whole Fc of human immunoglobulin E (hIgE-Fc) from mammalian cells at levels up to 100 mg/l of culture. Two surface glycosylation sites at Asn265 and Asn371 have been changed to glutamine, to obtain a more homogeneous preparation of hIgE-Fc for structural studies. Comparison of wild-type and mutant products revealed that Asn371 is rarely glycosylated in Chinese hamster ovary cells. Both the double mutant and wild-type hIgE-Fc bind to the high-affinity IgE receptor, Fc epsilon RI, with about the same affinity as myeloma IgE (Ka in the range 10(10)-10(11) M-1), and were able to sensitize isolated human basophils for anti-IgE triggering of histamine release. However, only the double mutant hIgE-Fc approached the affinity of myeloma IgE for the low-affinity receptor, Fc epsilon RII (Ka = 7.3 x 10(7) M-1), whereas the wild-type hIgE-Fc bound with a 10-fold lower affinity (Ka = 4.1 x 10(6) M-1).
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