Aim To investigate whether glycation could inhibit the protection role of mutant CD59s against human complement. Methods Site-directed mutagenesis to replace residue 37 or 38 with histidine(H)was performed by recombinant PCR.Mutant CD59 DNAs were inserted into the mammalian expression vector pALTER-MAX and transfected into CHO together with the selection marker pcDNA3,which confered resistance to G418.Expression of mutant CD59s in the G418-resistant clones were tested by Western blot,immunohistochemistry and FCM. A functional dye release assay was used to measure protection role of CD59s against human complement. Results Recombinant plasmids of pALTER-HM CD59 had been successfully constructed according to sequence and enzyme digestion analysis.Stable transfectants were selected by the addition of G418.Stable populations of CHO cell,which expressed relatively high levels of recombinant protein,were sorted by immunolabled technique.In Western blot assay,the mutant proteins expressed on CHO was about 20 kDa.Dye release assays confirmed both mutants still preserved CD59 activity of anticomplement,and glycation of CD59 in CHO increased their sensitivity to MAC-mediated lysis. Conclusions Residues by Site-directed mutagenesis to replace residue 37 or 38 with histidine still preserved CD59 activity of anticomplement.Mutant CD59s can be glycated in vitro and loses its most MAC-inhibitory function.The presence of this glycation motif in human mutant CD59,may help explain the distinct propensity of humans to develop vascular proliferative complications of diabetes.