title: Protein fibrillation lag times during kinetic inhibition.
abstract: Protein aggregation is linked to more than 30 human pathologies, including Alzheimer's and Parkinson's diseases. Since small oligomers that form at the beginning of the fibrillation process probably are the most toxic elements, therapeutic strategies involving fibril fragmentation could be detrimental. An alternative approach, named kinetic inhibition, aims to prevent fibril formation by using small ligands that stabilize the parent protein. The factors that govern fibrillation lag times during kinetic inhibition are largely unknown, notwithstanding their importance for designing effective long-term therapies. Inhibitor-bound species are not likely to be incorporated into the core of mature fibrils, although their presence could alter the kinetics of the fibrillation process. For instance, inhibitor-bound species may act as capping elements that impair the nucleation process and/or fibril growth. Here, we address this issue by studying the effect of two natural inhibitors on the fibrillation behavior of lysozyme at neutral pH. We analyzed a set of 79 fibrillation curves obtained in lysozyme alone and a set of 37 obtained in the presence of inhibitors. We calculated the concentrations of the relevant species at the beginning of the curves using the inhibitor-binding constants measured under the same experimental conditions. We found that inhibitor-bound protein species do not affect fibrillation onset times, which are mainly determined by the concentration of unbound protein species present in equilibrium. In this system, knowledge of the fibrillation kinetics and inhibitor affinities suffices to predict the effect of kinetic inhibitors on fibrillation lag times. In addition, we developed a new methodology to better estimate fibrillation lag times from experimental curves.
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina.
authors.forename: Rodrigo S
authors.initials: RS
authors.lastname: Pagano
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina.
authors.forename: Máximo
authors.initials: M
authors.lastname: López Medus
authors.affiliation: Department of Biological Chemistry, School of Pharmacy and Biochemistry, University of Buenos Aires, Buenos Aires, Argentina.
authors.forename: Gabriela E
authors.initials: GE
authors.lastname: Gómez
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina.
authors.forename: Paula M
authors.initials: PM
authors.lastname: Couto
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina.
authors.forename: María S
authors.initials: MS
authors.lastname: Labanda
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina.
authors.forename: Lucas
authors.initials: L
authors.lastname: Landolfo
authors.affiliation: Laboratory of Glycobiology, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina; Department of Physiology and Molecular Biology, University of Buenos Aires, Buenos Aires, Argentina.
authors.forename: Cecilia
authors.initials: C
authors.lastname: D'Alessio
authors.affiliation: Structural Cell Biology Laboratory, Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas de Buenos Aires (IIBBA-CONICET), Buenos Aires, Argentina; Department of Biological Chemistry, School of Sciences, University of Buenos Aires, Buenos Aires, Argentina. Electronic address: jcaramelo@leloir.org.ar.
authors.forename: Julio J
authors.initials: JJ
authors.lastname: Caramelo
mesh_terms: D000818:Animals; D002645:Chickens; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019791:Guanidine / Q000494:pharmacology; D007700:Kinetics; D009113:Muramidase / Q000037:antagonists & inhibitors / Q000737:chemistry*; D066329:Protein Aggregates
chemical_list: D004791:Enzyme Inhibitors; D066329:Protein Aggregates; D009113:Muramidase; D019791:Guanidine
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