Title : Disulfide bond structure and N-glycosylation
sites of the extracellular
domain of the human
interleukin-6 receptor
Abstract :
- The high affinity interleukin-6 ( IL-6 ) receptor is a hexameric complex consisting of two molecules each of IL-6 , IL-6 receptor ( IL-6R ), and the high affinity converter and signaling molecule, gp130
- The extracellular "soluble" part of the IL-6R ( sIL-6R) consists of three domains: an amino-terminal Ig-like domain and two fibronectin-type III (FN III) domains
- The two FN III domains comprise the cytokine-binding domain defined by a set of 4 conserved cysteine residues and a WSXWS sequence motif
- Here, we have determined the disulfide structure of the human sIL-6R by peptide mapping in the absence and presence of reducing agent
- Mass spectrometric analysis of these peptides revealed four disulfide bonds and two free cysteines
- The disulfides Cys102-Cys113 and Cys146-Cys157 are consistent with known cytokine-binding domain motifs , and Cys28-Cys77 with known Ig superfamily domains
- An unusual cysteine connectivity between Cys6-Cys174, which links the Ig-like and NH2-terminal FN III domains causing them to fold back onto each other, has not previously been observed among cytokine receptors
- The two free cysteines ( Cys192 and Cys258 ) were detected as cysteinyl- cysteines , although a small proportion of Cys258 was reactive with the alkylating agent 4-vinylpyridine
- Of the four potential N-glycosylation sites , carbohydrate moieties were identified on Asn36, Asn74, and Asn202 , but not on Asn226