Browsing by Author "Llauro, Aida"

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Changes in the stability and biomechanics of P22 bacteriophage capsid during maturation
(2018-06) Kant, Ravi; Llauro, Aida; Rayaprolu, Vamseedhar; Qazi, Shefah; de Pablo, Pedro J.; Douglas, Trevor; Bothner, Brian
The capsid of P22 bacteriophage undergoes a series of structural transitions during maturation that guide it from spherical to icosahedral morphology. The transitions include the release of scaffold proteins and capsid expansion. Although P22 maturation has been investigated for decades, a unified model that incorporates thermodynamic and biophysical analyses is not available. A general and specific model of icosahedral capsid maturation is of significant interest to theoreticians searching for fundamental principles as well as virologists and material scientists seeking to alter maturation to their advantage. To address this challenge, we have combined the results from orthogonal biophysical techniques including differential scanning fluorimetry, atomic force microscopy, circular dichroism, and hydrogen-deuterium exchange mass spectrometry. By integrating these results from single particle and population measurements, an energy landscape of P22 maturation from procapsid through expanded shell to wiffle ball emerged, highlighting the role of metastable structures and the thermodynamics guiding maturation. The propagation of weak quaternary interactions across symmetric elements of the capsid is a key component for stability in P22. A surprising finding is that the progression to wiffle ball, which lacks pentamers, shows that chemical and thermal stability can be uncoupled from mechanical rigidity, elegantly demonstrating the complexity inherent in capsid protein interactions and the emergent properties that can arise from icosahedral symmetry. On a broader scale, this work demonstrates the power of applying orthogonal biophysical techniques to elucidate assembly mechanisms for supramolecular complexes and provides a framework within which other viral systems can be compared.
Decrease in pH destabilizes individual vault nanocages by weakening the inter-protein lateral interaction
(2016-10) Llauro, Aida; Guerra, Pablo; Kant, Ravi; Bothner, Brian; Verdaguer, Nuria; de Pablo, Pedro J
Vault particles are naturally occurring proteinaceous cages with promising application as molecular containers. The use of vaults as functional transporters requires a profound understanding of their structural stability to guarantee the protection and controlled payload delivery. Previous results performed with bulk techniques or at non-physiological conditions have suggested pH as a parameter to control vault dynamics. Here we use Atomic Force Microscopy (AFM) to monitor the structural evolution of individual vault particles while changing the pH in real time. Our experiments show that decreasing the pH of the solution destabilize the barrel region, the central part of vault particles, and leads to the aggregation of the cages. Additional analyses using Quartz-Crystal Microbalance (QCM) and Differential Scanning Fluorimetry (DSF) are consistent with our single molecule AFM experiments. The observed topographical defects suggest that low pH weakens the bonds between adjacent proteins. We hypothesize that the observed effects are related to the strong polar character of the protein-protein lateral interactions. Overall, our study unveils the mechanism for the influence of a biologically relevant range of pHs on the stability and dynamics of vault particles.