RESUMEN
The deposition of pathological protein aggregates in the brain plays a central role in cognitive decline and structural damage associated with neurodegenerative diseases. In Alzheimer's disease, the formation of amyloid-ß plaques and neurofibrillary tangles of the tau protein is associated with the appearance of symptoms and pathology. Detailed models for the specific mechanisms of aggregate formation, such as nucleation and elongation, exist for aggregation in vitro where the total protein mass is conserved. However, in vivo, an additional class of mechanisms that clear pathological species is present and is believed to play an essential role in limiting the formation of aggregates and preventing or delaying the emergence of disease. A key unanswered question in the field of neuro-degeneration is how these clearance mechanisms can be modeled and how alterations in the processes of clearance or aggregation affect the stability of the system toward aggregation. Here, we generalize classical models of protein aggregation to take into account both production of monomers and the clearance of protein aggregates. We show that, depending on the specifics of the clearance process, a critical clearance value emerges above which accumulation of aggregates does not take place. Our results show that a sudden switch from a healthy to a disease state can be caused by small variations in the efficiency of the clearance process and provide a mathematical framework to explore the detailed effects of different mechanisms of clearance on the accumulation of aggregates.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Enfermedades Neurodegenerativas/metabolismo , Agregación Patológica de Proteínas/metabolismo , Péptidos beta-Amiloides/química , Humanos , Cinética , Modelos QuímicosRESUMEN
While a significant body of investigations have been focused on the process of protein self-assembly, much less is understood about the reverse process of a filament breaking due to thermal motion into smaller fragments, or depolymerization of subunits from the filament ends. Indirect evidence for actin and amyloid filament fragmentation has been reported, although the phenomenon has never been directly observed either experimentally or in simulations. Here we report the direct observation of filament depolymerization and breakup in a minimal, calibrated model of coarse-grained molecular simulation. We quantify the orders of magnitude by which the depolymerization rate from the filament ends koff is larger than fragmentation rate k- and establish the law koff/k- = exp[(εâ - εâ¥)/kBT] = exp[0.5ε/kBT], which accounts for the topology and energy of bonds holding the filament together. This mechanism and the order-of-magnitude predictions are well supported by direct experimental measurements of depolymerization of insulin amyloid filaments.
Asunto(s)
Actinas/química , Amiloide/química , Modelos Moleculares , Multimerización de Proteína , Actinas/metabolismo , Amiloide/metabolismo , Cinética , Estructura Secundaria de Proteína , TemperaturaRESUMEN
Micro- and nano-scale systems have emerged as important tools for developing clinically useful drug delivery systems. In this tutorial review, we discuss the exploitation of biomacromolecules for this purpose, focusing on proteins, polypeptides, nucleic acids and polysaccharides and mixtures thereof as potential building blocks for novel drug delivery systems. We focus on the mechanisms of formation of micro- and nano-scale protein-based capsules and shells, as well as on the functionalization of such structures for use in targeted delivery of bioactive materials. We summarise existing methods for protein-based capsule synthesis and functionalization and highlight future challenges and opportunities for delivery strategies based on biomacromolecules.
Asunto(s)
Portadores de Fármacos/química , Nanocápsulas/química , Proteínas/química , Tamaño de la Partícula , Estabilidad Proteica , Proteínas/metabolismoRESUMEN
We consider the spatial dependence of filamentous protein self-assembly. Through studying the cases where the spreading of aggregated material is dominated either by diffusion or by growth, we derive analytical results for the spatial evolution of filamentous protein aggregation, which we validate against Monte Carlo simulations. Moreover, we compare the predictions of our theory with experimental measurements of two systems for which we identify the propagation as either growth or diffusion controlled. Our results connect the macroscopic observables that characterize the spatial propagation of protein self-assembly with the underlying microscopic processes and provide physical limits on spatial propagation and prionlike behavior associated with protein aggregation.
Asunto(s)
Modelos Químicos , Proteínas/química , Difusión , Método de Montecarlo , Polimerizacion , Proteínas/metabolismo , Procesos EstocásticosRESUMEN
As the use of microfluidic chips for handling biological samples is increasing, so is the need for combining them with powerful analytical techniques for metal determination such as inductively-coupled plasma mass spectrometry (ICP-MS). So far, coupling a microfluidic chip to an ICP-MS has been demonstrated mainly through the use of conventional pneumatic micro-flow nebulizers. However, disadvantages associated with the use of such nebulizers entail dead volume issues and liquid suction exerted on the outlet channel of the chip. Herein, we propose that a microfluidic chip, bearing a pneumatic nozzle for liquid nebulization, has the potential to advance metal determination in chip-based ICP-MS. More specifically, we demonstrate for the first time that the coupling of a chip-based supersonic microfluidic nebulizer (chip-µf-Neb) to an ICP-MS can be conveniently achieved through the use of a spray chamber with a laminar flow makeup gas. Operation of the combined system was evaluated at low liquid flow rates across 0.5-20 µL min-1, while nebulization and makeup argon (Ar) gas flow rates were optimized with respect to maximizing indium (In) sensitivity and minimizing oxide formation; a maximum sensitivity of 40000 cps (µg L-1)-1 was achieved at 10 µL min-1. The system was further evaluated for its performance in single-particle analysis, featuring a transport efficiency of 46% for Ag nanoparticles. Finally, the capabilities for conducting single-cell analysis were demonstrated with the detection of 80Se and 75As in individual Chlamydomas reinhardtii cells, which were previously incubated in 20 µM of selenate and 300 µM of arsenate, respectively. Efficient operation at low liquid flow rates along with the absence of self-aspiration render this nebulizer a promising tool for combining the powerful field of microfluidics with metal quantitation by means of ICP-MS.
Asunto(s)
Nanopartículas del Metal , Microfluídica , Argón , Arseniatos , Indio , Espectrometría de Masas/métodos , Nebulizadores y Vaporizadores , Óxidos , Ácido Selénico , PlataRESUMEN
Cell membranes are integral to the functioning of the cell and are therefore key to drive fundamental understanding of biological processes for downstream applications. Here, we review the current state-of-the-art with respect to biomembrane systems and electronic substrates, with a view of how the field has evolved towards creating biomimetic conditions and improving detection sensitivity. Of particular interest are conducting polymers, a class of electroactive polymers, which have the potential to create the next step-change for bioelectronics devices. Lastly, we discuss the impact these types of devices could have for biomedical applications.
Asunto(s)
Técnicas Biosensibles , Electrónica , Biomimética , Técnicas Biosensibles/métodos , Membrana Celular , PolímerosRESUMEN
The composition of soluble toxic protein aggregates formed in vivo is currently unknown in neurodegenerative diseases, due to their ultra-low concentration in human biofluids and their high degree of heterogeneity. Here we report a method to capture amyloid-containing aggregates in human biofluids in an unbiased way, a process we name amyloid precipitation. We use a structure-specific chemical dimer, a Y-shaped, bio-inspired small molecule with two capture groups, for amyloid precipitation to increase affinity. Our capture molecule for amyloid precipitation (CAP-1) consists of a derivative of Pittsburgh Compound B (dimer) to target the cross ß-sheets of amyloids and a biotin moiety for surface immobilization. By coupling CAP-1 to magnetic beads, we demonstrate that we can target the amyloid structure of all protein aggregates present in human cerebrospinal fluid, isolate them for analysis and then characterize them using single-molecule fluorescence imaging and mass spectrometry. Amyloid precipitation enables unbiased determination of the molecular composition and structural features of the in vivo aggregates formed in neurodegenerative diseases.