Association of XK and Kell blood group proteins

A disulfide bond links Kell and XK red cell membrane proteins. Kell, a type II membrane glycoprotein, carries over 20 blood group antigens, and XK, which spans the membrane 10 times, is lacking in rare individuals with the McLeod syndrome. Kell is classified in the neprilysin family of zinc endopeptidases, and XK has structural features that suggest it is a transport protein. Kell has 15 extracellular cysteines, and XK has one in its fifth extracellular loop. Five of the extracellular cysteine residues in Kell are not conserved in the other members of the neprilysin family, and based on the hypothesis that one of the nonconserved cysteines is linked to XK, cysteines 72 and 319 were mutated to serine. The single extracellular cysteine 347 of XK was also mutated. Co-expression of combinations of wild-type and mutant proteins in transfected COS-1 cells showed that Kell C72S did not form a Kell-XK complex with wild-type XK, while wild-type Kell and Kell C319S did. XK C347S was also unable to form a complex with wild-type Kell, indicating that Kell cysteine 72 is linked to XK cysteine 347. Kell C72S was transported to the cell surface, indicating that linkage to XK is not required. In addition, chemical cross-linking of red cell membranes with dithiobispropionimidate indicated that glyceraldehyde-3-phosphate dehydrogenase is a near neighbor of Kell.