Idioma:

19F NMR Studies of Plasminogen Activator Inhibitor-1

Abbott, Glenn L ; Blouse, Grant E ; Perron, Michel J ; Shore, Joseph D ; Luck, Linda A ; Szabo, Arthur G

Biochemistry (Easton), 2004-02, Vol.43 (6), p.1507-1519 [Revista revisada por pares]

United States: American Chemical Society

Texto completo disponible

Citas Citado por

Acciones

Título:
19F NMR Studies of Plasminogen Activator Inhibitor-1
Autor: Abbott, Glenn L ; Blouse, Grant E ; Perron, Michel J ; Shore, Joseph D ; Luck, Linda A ; Szabo, Arthur G
Materias: Fluorine - chemistry ; Humans ; Hydrolysis ; Mutagenesis, Site-Directed ; Nuclear Magnetic Resonance, Biomolecular ; Plasminogen Activator Inhibitor 1 - biosynthesis ; Plasminogen Activator Inhibitor 1 - chemistry ; Plasminogen Activator Inhibitor 1 - genetics ; Recombinant Proteins - biosynthesis ; Recombinant Proteins - chemistry ; Recombinant Proteins - genetics ; Serine Proteinase Inhibitors - chemistry ; Serine Proteinase Inhibitors - genetics ; Structure-Activity Relationship ; Tissue Plasminogen Activator - antagonists & inhibitors ; Tissue Plasminogen Activator - chemistry ; Tryptophan - analogs & derivatives ; Tryptophan - chemistry ; Tryptophan - genetics
Es parte de: Biochemistry (Easton), 2004-02, Vol.43 (6), p.1507-1519
Notas: ark:/67375/TPS-X84VPWLW-2
This work was supported by the Heart and Stroke Foundation of Ontario. L.A.L. acknowledges partial funding for this work from the Petroleum Research Fund (Grant 36825-AC4).
istex:66B085B4D4BB430332F55B8F9F6BA39FAC7E2C72
Descripción: Plasminogen activator inhibitor-1 (PAI-1) is a 43 kDa protein involved in the regulation of fibrinolysis. PAI-1 is the principal inhibitor of tissue-type plasminogen activator (t-PA), trapping the proteinase as an acyl-enzyme covalent complex (∼105 kDa). Four single tryptophan mutants of PAI-1 have been constructed in which three of the four tryptophan residues (Trp86, Trp139, Trp175, and Trp262) were replaced with phenylalanine. Biosynthetic incorporation of 5-fluorotryptophan (5F-Trp) into wild-type PAI-1 (5FW wtPAI-1) and the single tryptophan mutants (5FW86, 5FW139, 5FW175, and 5FW262) was achieved, allowing a 19F NMR spectroscopic study of PAI-1 in its active and cleaved forms and in complex with t-PA. The 19F NMR spectrum of active 5FW wtPAI-1 shows four clearly resolved peaks at −39.20, −49.26, −50.74, and −52.57 ppm relative to trifluoroacetic acid at 0 ppm. Unequivocal assignments of these four resonances in the spectrum of 5FW wtPAI-1 to specific tryptophan residues were accomplished by measuring the chemical shifts of the 19F resonances of the single tryptophan mutants. There was close agreement between the resonances observed in 5FW wtPAI-1 and of those in the mutants for all three protein forms. This would imply little structural perturbation in the local structures of the tryptophan residues resulting from substitution by phenylalanine. The 5FW wtPAI-1 was observed to have lower second-order rate constant (k app) for the inhibition of t-PA than the natural tryptophan wtPAI-1, suggesting that the decreased activity may result from a small structural effect of the fluorine substituent of the indole ring. Further alterations in the k app and the stoichiometry of inhibition (SI) were observed in each of the mutants indicating an effect of the three tryptophan to phenylalanine mutations. Detailed interpretation of the 19F NMR spectra of the PAI-1 mutants provides insights into the local segmental structure of the active form of the proteins and the structural changes that occur in the cleaved and t-PA complexed forms.
Editor: United States: American Chemical Society
Idioma: Inglés

Enlaces

View this record in MEDLINE/PubMed

Volver a la lista de resultados

Anterior Resultado 5 Siguiente Ir a la siguiente página

Realización: Logos de Redes Sociales:

19F NMR Studies of Plasminogen Activator Inhibitor-1

Abbott, Glenn L ; Blouse, Grant E ; Perron, Michel J ; Shore, Joseph D ; Luck, Linda A ; Szabo, Arthur G

United States: American Chemical Society

Buscando en bases de datos remotas, por favor espere