Secondary amyloid A (AA) amyloidosis is an important complication of some chronic inflammatory diseases, primarily rheumatoid arthritis (RA). It is a serious, potentially life-threatening disorder caused by the deposition of AA fibrils, which are derived from the circulatory, acute-phase-reactant, serum amyloid A protein (SAA). Recently, a specific interaction between SAA and the ubiquitous inhibitor of cysteine proteases--human cystatin C (hCC)--has been proved. Using a combination of selective proteolytic excision and high-resolution mass spectrometry, the binding sites in the SAA and hCC sequences were assessed as SAA(86-104) and hCC(96-102), respectively. Here, we report further details concerning the hCC-SAA interaction. With the use of affinity tests and florescent ELISA-like assays, the amino acid residues crucial for the protein interaction were determined. It was shown that all amino acid residues in the SAA sequence, essential for the formation of the protein complex, are basic ones, which suggests an electrostatic interaction character. The idea is corroborated by the fact that the most important residues in the hCC sequence are Ser-98 and Tyr-102; these residues are able to form hydrogen bonds via their hydroxyl groups. The molecular details of hCC-SAA complex formation might be helpful for the design of new compounds modulating the biological role of both proteins.
Keywords: Serum amyloid A; binding sites; human cystatin C; mass spectrometry.
Copyright © 2013 John Wiley & Sons, Ltd.