Sökning: id:"swepub:oai:DiVA.org:umu-8572" >
The structure of a ...
The structure of a serpin–protease complex revealed by intramolecular distance measurements using donor–donor energy migration and mapping of interaction sites
-
- Fa, Ming (författare)
- Umeå universitet,Institutionen för medicinsk kemi och biofysik
-
- Bergström, Fredrik (författare)
- Umeå universitet,Kemiska institutionen
-
- Hägglöf, Peter (författare)
- Umeå universitet,Institutionen för medicinsk kemi och biofysik
-
visa fler...
-
- Wilczynska, Malgorzata (författare)
- Umeå universitet,Institutionen för medicinsk kemi och biofysik
-
- Johansson, Lennart B-Å (författare)
- Umeå universitet,Kemiska institutionen
-
- Ny, Tor (författare)
- Umeå universitet,Institutionen för medicinsk kemi och biofysik
-
visa färre...
-
(creator_code:org_t)
- 2000
- 2000
- Engelska.
-
Ingår i: Structure. ; 8:4, s. 397-405
- Relaterad länk:
-
https://urn.kb.se/re...
-
visa fler...
-
https://doi.org/10.1...
-
visa färre...
Abstract
Ämnesord
Stäng
- Background: The inhibitors that belong to the serpin family are widely distributed regulatory molecules that include most protease inhibitors found in blood. It is generally thought that serpin inhibition involves reactive-centre cleavage, loop insertion and protease translocation, but different models of the serpin–protease complex have been proposed. In the absence of a spatial structure of a serpin–protease complex, a detailed understanding of serpin inhibition and the character of the virtually irreversible complex have remained controversial.Results: We used a recently developed method for making precise distance measurements, based on donor–donor energy migration (DDEM), to accurately triangulate the position of the protease urokinase-type plasminogen activator (uPA) in complex with the serpin plasminogen activator inhibitor type 1 (PAI-1). The distances from residue 344 (P3) in the reactive-centre loop of PAI-1 to residues 185, 266, 313 and 347 (P1′) were determined. Modelling of the complex using this distance information unequivocally placed residue 344 in a position at the distal end from the initial docking site with the reactive-centre loop fully inserted into β sheet A. To validate the model, seven single cysteine substitution mutants of PAI-1 were used to map sites of protease–inhibitor interaction by fluorescence depolarisation measurements of fluorophores attached to these residues and cross-linking using a sulphydryl-specific cross-linker.Conclusions: The data clearly demonstrate that serpin inhibition involves reactive-centre cleavage followed by full-loop insertion whereby the covalently linked protease is translocated from one pole of the inhibitor to the opposite one.
Nyckelord
- Cross-linking
- Donor–donor energy migration
- Fluorescence
- Intramolecular distance
- PAI-1
- serpin
Publikations- och innehållstyp
- ref (ämneskategori)
- art (ämneskategori)
Hitta via bibliotek
Till lärosätets databas