title: Interaction of serum amyloid A with human cystatin C--identification of binding sites.
abstract: Serum amyloid A (SAA) is a multifunctional acute-phase protein whose natural role seems to be participation in many physiologic and pathological processes. Prolonged increased SAA level in a number of chronic inflammatory and neoplastic diseases gives rise to reactive systemic amyloid A amyloidosis, where the N-terminal 76-amino acid residue-long segment of SAA is deposited as amyloid fibrils. Recently, a specific interaction between SAA and the ubiquitous inhibitor of cysteine proteases--human cystatin C (hCC)--has been described. Here, we report further evidence corroborating this interaction, and the identification of the SAA and hCC binding sites in the SAA-hCC complex, using a combination of selective proteolytic excision and high-resolution mass spectrometry. The shortest binding site in the SAA sequence was determined as SAA(86-104), whereas the binding site in hCC sequence was identified as hCC(96-102). Binding specificities of both interacting sequences were ascertained by affinity experiments (ELISA) and by registration of mass spectrum of SAA-hCC complex.
authors.affiliation: Department of Medicinal Chemistry, University of Gdansk, Sobieskiego 18, 80-952 Gdansk, Poland.
authors.forename: Marta
authors.initials: M
authors.lastname: Spodzieja
authors.forename: Aneta
authors.initials: A
authors.lastname: Szymańska
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Kołodziejczyk
authors.forename: Martyna
authors.initials: M
authors.lastname: Prądzińska
authors.forename: Martyna
authors.initials: M
authors.lastname: Maszota
authors.forename: Piotr
authors.initials: P
authors.lastname: Stefanowicz
authors.forename: Zbigniew
authors.initials: Z
authors.lastname: Szewczuk
authors.forename: Anders
authors.initials: A
authors.lastname: Grubb
authors.forename: Paulina
authors.initials: P
authors.lastname: Czaplewska
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D055316:Cystatin C / Q000737:chemistry* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D055767:Immobilized Proteins / Q000737:chemistry / Q000235:genetics; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D059748:Proteolysis; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D000685:Serum Amyloid A Protein / Q000737:chemistry* / Q000235:genetics; D012996:Solutions
chemical_list: D055316:Cystatin C; D055767:Immobilized Proteins; D011994:Recombinant Proteins; D000685:Serum Amyloid A Protein; D012996:Solutions