Document details

Dynamic and interaction of cytochrome c with Pf1 virus

Author(s): Nunes, Patrique Nelson Ramos cv logo 1

Date: 2011

Persistent ID: http://hdl.handle.net/10362/5662

Origin: Repositório Institucional da UNL

Subject(s): Cytochrome c; Pf1 virus; Stoichiometry complex; Electrostatic interaction; Ionic strength; NMR-DOSY


Description
Cytochrome c is a positive protein and the Pf1 virus surface is negative forging strong electrostatic complex. When a critical ratio concentration of Cytochrome c and Pf1 virus is achieved a spontaneous complex is formed. The maximum association upon addition of cytochrome c to Pf1 solutions is about 1700 cytochrome c molecules to one Pf1 virion particle. The effect of univalent salt concentration on protein polyelectrolyte complex formation was measured by Dynamic Light Scattering. Complex disaggregation occurred when monovalent salt concentration increased. The assembly process was also observed by NMR at low salt concentration in the system. The aggregate can be gradually dissociated in order to enable NMR spectra acquisition. Depending on virus/cytochorme c ratio or ionic strength concentration we could shift from free protein and virus in solution to transient binding or fully immobilized complex. It was possible to map the most affected regions of the oxidized heme cytochrome c, with chemical shift variation due to the binding to Pf1 virus, during salt titration. Dry and liquid samples of Cytochrome c and Pf1 at different ratios and pH were studied and evaluated by Atomic Force Microscopy. The system was also studied above the critical salt concentration of complex dissociation by PGSEDOSY NMR. A gradual decrease in the translational diffusion coefficient of cytochrome was caused by higher content of Pf1 virus in solution. We conclude that a strong electrostatic correlation between pf1 virus and cytochrome c occurs even after complex dissociation. Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em BioOrgânica
Document Type Master Thesis
Language English
Advisor(s) Caldeira, Francisco
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