Cookies on this website

We use cookies to ensure that we give you the best experience on our website. If you click 'Accept all cookies' we'll assume that you are happy to receive all cookies and you won't see this message again. If you click 'Reject all non-essential cookies' only necessary cookies providing core functionality such as security, network management, and accessibility will be enabled. Click 'Find out more' for information on how to change your cookie settings.

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).

Type

Journal article

Journal

Protein Eng

Publication Date

02/1995

Volume

8

Pages

193 - 199

Keywords

Amino Acid Sequence, Animals, Base Sequence, CHO Cells, Cricetinae, Deoxyribonuclease EcoRI, Deoxyribonucleases, Type II Site-Specific, Electrophoresis, Polyacrylamide Gel, Glycosylation, Histamine Release, Humans, Immunoglobulin E, Immunoglobulin Fc Fragments, Macromolecular Substances, Molecular Sequence Data, Mutagenesis, Site-Directed, Recombinant Proteins, Structure-Activity Relationship, Transfection