[Study on mutation, expression and activity of human CD59 gene].
OBJECTIVE
To construct an eukaryotic expression system of human mutant CD59 (hmCD59) gene, and investigate the effect of hmCD59 on diabetic vascular complications.


METHODS
Two recombinant pALTER plasmids containing two hmCD59 cDNAs were transfected into CHO cells respectively with pcDNA3 by lipofectamine. Stably transfected clones were then selected by G418. Expression of hmCD59 on CHO cells was detected by fluorescent antibody technique, immunoenzymatic histochemistry and Western blot. 2', 7'-bis(carboxyethyl)-5, 6-carboxyfluorescein acetoxymethylester (BCECF/AM) was used to study the anti-complement activity of hmCD59 before and after glycosylation.


RESULTS
hmCD59 was expressed on stably transfected CHO cell surface. Both hmCD59 had the anti-complement activity which was weakened after glycosylation.


CONCLUSIONS
Two stable cell lines expressing hmCD59 have been successfully established. Both hmCD59 had the anti-complement activity, which was weakened after glycosylation.
Zhang
Yan-li
YL
Department of Immunology, Qingdao University Medical College, Qingdao 266021, China. ZZYYLL88@yahoo.com
Gao
Mei-hua
MH
D000818:Animals; D018957:CD59 Antigens / Q000235:genetics* / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003925:Diabetic Angiopathies / Q000175:diagnosis / Q000378:metabolism; D015870:Gene Expression; D006031:Glycosylation; D006801:Humans; D050505:Mutant Proteins / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D014162:Transfection
D018957:CD59 Antigens; D050505:Mutant Proteins