Servicio de Información Comunitario sobre Investigación y Desarrollo - CORDIS


MOLRHEOSTAT Informe resumido

Project ID: 323059
Financiado con arreglo a: FP7-IDEAS-ERC
País: Spain

Mid-Term Report Summary - MOLRHEOSTAT (Downhill Folding Protein Modules as Conformational Rheostats: Roles in Molecular Biology and Applications as Biosensors)

MOLRHEOSTAT aims at growing the grassroots of quantitative and synthetic molecular biology by focusing on the investigation of novel connections between protein folding and function via a multidisciplinary, holistic approach that combines experiment, theory and computation. Conventionally, proteins are portrayed as conformational switches that fold and function by toggling between an on-state (native, active) and an off-state (inactive, unfolded) in response to stimuli. However, over the last years we have witnessed the discovery of protein modules that undergo continuous conformational changes upon unfolding (downhill folding). Our overarching goal is to determine whether downhill folding modules operate mechanistically as conformational rheostats; that is, as single-molecular devices able to continuously modulate signals or responses by gradually tuning their folding conformational ensemble. Conformational rheostats have strategic interest because they would open a new realm of functional opportunities for controlling complex biochemical processes, and could have tremendous technological impact as biomolecular scaffolds for building synthetic nanodevices. Our group pioneered the experimental characterization of downhill folding modules and the analysis of their remarkable conformational behavior through combined advanced biophysical spectroscopic methods and theoretical approaches. Now we are using our expertise accumulated over 15 years of work in this area to explore their functional roles and technological potential via a research plan divided into two specific and complementary objectives: 1) implementation of a general approach for building high-performance, ultrafast, single-molecule sensors based on downhill folding modules; 2) analysis of the roles of conformational rheostats in the regulation of several fundamental processes in molecular biology.

In objective 1 we are building nanoscale biosensors based on the gradual coupling between downhill folding and binding to transduce analogical signals at the single-molecule level. By effectively engineering proof-of-concept sensing nanodevices to measure pH and ionic strength we are taking important strides in demonstrating the practical feasibility of the conformational rheostat concept. We are also developing a biomolecular engineering toolset to facilitate the implementation of rheostat-based high-performance single-molecule nanosensors. Such toolbox includes computational methods for identifying new downhill modules, protein engineering protocols for manipulating the folding mechanism and stability of target proteins, and strategies for implementing gradual fluorescence readouts.

Our work towards objective 2 is based on the premise that Nature has exploited the conformational rheostat mechanism on key biological processes that require analogic control at the single molecule level. To investigate this general hypothesis we are investigating three fundamental biological processes that involve multivariate molecular functions carried out by downhill folding modules: 1) molecular hubs in protein-protein interaction network that exert control by binding to multiple, structurally diverse protein partners via conformational selection and induced-fit; 2) the conformational mechanisms that allow DNA-binding domains to slide over DNA, hop and home into the specific target sequence; 3) the mechanical roles of downhill folding modules as molecular springs in macromolecular assemblies.

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