NMR structure of the S-domain of calreticulin, the sub-fragment that comprises the calreticulin binding site for defence collagens

• Main Author: Gingras, Alexandre R.
• Published: University of Leicester 2004
• Subjects: 572.69
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.483577

Using the overlapping sub-fragments, i.e. P-, S- and SP-domains of CRT in different binding assays, we determined that the S-domain has the strongest affinity to bind to C1q and MBL. This makes the structure of the S-domain most relevant for understanding the binding interactions to C1q and MBL and the sterical inhibition of the complement activation by competing with the respective C1q or MBL associated serine proteases for binding. The CRT S-domain structure determined in this work has an extended hairpin fold stabilised by a short 3 residues beta-sheet holding the two opposite strands together. Two hydrophobic cores, each containing two conserved tryptophan residues, are located on either sides of this beta-sheet and further stabilise this fold. Comparison of 2D [1H, 15N]-HSQC spectra indicated higher internal mobility of the S-domain than of the other active fragments, suggesting that the dynamical properties are important for the protein activity. Detailed multi-field relaxation analysis showed high dynamics in the hairpin with more restrictions where secondary structure elements are present. The interaction with MBL and C1q was determined to be optimal under low ionic strength conditions, indicative of electrostatic interactions.;The interaction probably involves the tip of the hairpin in the CRT S-domain structure that has a high concentration of negatively charged residues, and a complimentary positively charged region of the MBL and C1q collagen-like domains. The flexibility of the hairpin is likely to be a key element for CRT, allowing it to adopt its conformation for binding to multiple ligands and is expected to be the key factor for the enhanced activity of the S-domain.