Bridging scales in the coevolution of interacting proteins
Martin Weigt (Institut de biologie Paris-Seine)
Understanding protein−protein interactions (PPI) is central to our understanding of almost all complex biological processes. Computational tools exploiting rapidly growing genomic databases to characterize PPI are therefore urgently needed. Such methods should address multiple scales of PPI: (i) Between two interacting proteins, which residues are in contact across the interfaces? (ii) Inside a genome, which specific proteins interact and which do not? (iii) On evolutionary time scales, which protein-protein interactions are actually conserved across thousands of species? Statistical inference methods like the Direct-Coupling Analysis (DCA), have recently triggered considerable progress in using sequence data to connect these different scales, thereby helping to assemble quaternary protein structures and to predict conserved interactions between proteins. Besides evident bioinformatic applications in structural and systems biology, these methods help to deepen our understanding of the patterns of co-evolution between interacting proteins in general.
Liquid crystal ordering of RNA: a route for the origin of life?
We recently discovered that concentrated solutions of short oligonucleotides (down to 4 base long) and even solutions of single bases spontaneously develop liquid crystal ordering.
In such supramolecular assemblies, duplex-forming oligomers and paired bases are held in continuous mutual contact to form chemically discontinuous but physically continuous double helices.
We found that this spontaneous order can serve as a mechanism for molecular selection and as a template to guide abiotic ligation of the oligomers in long chains.
Specifically, we studied the influence of liquid crystal ordering on the efficiency of non-enzymatic ligation reaction induced by water-soluble carbodiimide EDC as condensing agent.
We find that the liquid crystal ordering markedly enhances the ligation efficacy, thus suggesting such spontaneous ordering may have been the key feature in the emerging of nucleic acids from the primordial molecular noise.
Was the RNA World prepared by an even more ancient “Liquid Crystal World”?
Bridging length and time scales in filamentous protein self-assembly
Filamentous protein self-assembly is a process in which dispersed proteins assemble spontaneously to form ordered elongated structures. This phenomenon is an essential characteristic of life, but is also at the heart of pathologies of many types, including Parkinson’s and Alzheimer’s diseases. Moreover, filamentous growth is increasingly used in numerous nanotechnological applications. To exploit filamentous self-assembly for nanotechnology or curtail it for medical purposes it is necessary to quantify the fundamental principles that control the way dispersed molecules assemble into these ordered structures. The fundamental challenge in establishing such an understanding in a rigorous manner is the disparate nature of the spatial and temporal scales involved, which range from the molecular to the organism scale. In this talk, I demonstrate how this challenge can be partially addressed by bringing the power of physical methods to protein filament self-assembly to connect microscopic mechanisms with macroscopic observations of such phenomena. In a first part of the talk, I discuss a unified theory of the kinetics of filamentous protein assembly and show how these results reveal simple rate laws that provide the basis for interpreting experimental data in terms of specific mechanisms controlling the proliferation of fibrils. I also describe methods for understanding the factors that control the size of linear aggregating proteins, a crucial parameter that correlates with their neurotoxicity, and discuss how this framework allows us to study the modes of action through which molecular chaperones can suppress amyloid fibril formation. In a second part of the talk, I investigate filamentous growth reactions under confinement where statistical mechanical fluctuations play a significant role and illustrate how this strategy can be used to establish further mechanistic insights into protein filament formation. Finally, I explore the possibilities and performance limits of force generation and energy release by nanoscale self-assembling supra-molecular polymers.
Snapshots on secretion, folding, translocation and functions of the Bordetella pertussis CyaA toxin
The adenylate cyclase toxin (CyaA, 1706 residues) plays an essential role in the early stages of respiratory tract colonization by Bordetella pertussis, the causative agent of whooping cough. The cell intoxication process, which is still poorly understood, involves a unique mechanism of translocation of the CyaA catalytic domain (AC) directly across the plasma membrane of target cells. Once in the cytosol, AC interacts with calmodulin (CaM) and produces supraphysiological levels of cAMP, leading to cell death. I will present some recent results covering several steps of the intoxication process, from toxin secretion to the AC:CaM complex formation. Our data illustrate the structural flexibility of bacterial toxins adapted to various functions (such as secretion and enzymatic complex formation) and coupled to large variations in calcium concentrations encountered in the successive environments during the intoxication process. Moreover, we recently showed that the translocation process is dependent on (i) the electrochemical gradient across the membrane and (ii) the unique properties of a short CyaA region that mimics membrane-active peptides. Finally, due to its hydrophobic character, CyaA toxins do aggregate into multimeric forms in the absence of chaotropic agents in vitro. We have recently defined the experimental conditions required for CyaA folding into a stable, monomeric and functional form. This opens new perspectives for both basic sciences and CyaA-based biotechnological applications developed in the lab, i.e., to improve antigen delivery vehicles and new pertussis vaccines.
An evolutionary switch in J-protein biology affects prokaryotic and eukaryotic protein disaggregation
Decreased cellular capacity in protein aggregate clearance manifests in cellular deterioration, aging and disease1. Toxic intracellular aggregates formed by misfolded proteins are reversed and/or limited by multi-tiered cellular quality control systems2. We recently reported J-proteins of classes A and B cooperate via interclass complex formation to mediate substrate specificity of Hsp70-based aggregate solubilizing systems (disaggregases) in metazoa3. What remains unclear is whether these mixed class J-protein complexes occur also in non-metazoans given the fact that orthologs of both classes exist in bacteria, fungi, plant and protozoa4. Using a broad set of experimental approaches, we find a switching in J-protein biology at the prokaryote-to-eukaryote transition where class members network allowing for the emergence of powerful, yet regulatable eukaryotic disaggregase systems. We also describe a naturally occurring strategy to correctly pair J-proteins of different types, ensuing functional integrity within networks in expanded J-protein families during rise of complex life.