The human [alpha]2-plasmin inhibitor: functional characterization of the unique plasmin(ogen)-binding region

• Published in: Cellular and molecular life sciences : CMLS 2010-05, Vol.67 (9), p.1505
• Main Authors: Gerber, Simon S, Lejon, Sofia, Locher, Michael, Schaller, Johann
• Format: Article
• Language: English
• Subjects: Binding sites; Cellular biology; E coli; Inhibitor drugs; Molecular biology; Mutation
• ISSN: 1420-682X; 1420-9071
• DOI: 10.1007/s00018-010-0264-3

The human [alpha]^sub 2^-plasmin inhibitor (A2PI) possesses unique N- and C-terminal extensions that significantly influence its biological activities. The C-terminal segment, A2PIC (Asn^sup 398^-Lys^sup 452^), contains six lysines thought to be involved in the binding to lysine-binding sites in the kringle domains of human plasminogen, of which four (Lys^sup 422^, Lys^sup 429^, Lys^sup 436^, Lys^sup 452^) are completely and two (Lys^sup 406^, Lys^sup 415^) are partially conserved. Multiple Lys to Ala mutants of A2PIC were expressed in Escherichia coli and used in intrinsic fluorescence titrations with kringle domains K1, K4, K4 + 5, and K1 + 2 + 3 of human plasminogen. We were able to identify the C-terminal Lys^sup 452^ as the main binding partner in recombinant A2PIC (rA2PIC) constructs with isolated kringles. We could show a cooperative, zipper-like enhancement of the interaction between C-terminal Lys^sup 452^ and internal Lys^sup 436^ of rA2PIC and isolated K1 + 2 + 3, whereas the other internal lysine residues contribute only to a minor extent to the binding process. Sulfated Tyr^sup 445^ in the unique C-terminal segment revealed no influence on the binding affinity to kringle domains. [PUBLICATION ABSTRACT]