27,936 research outputs found
A correspondence between solution-state dynamics of an individual protein and the sequence and conformational diversity of its family.
Conformational ensembles are increasingly recognized as a useful representation to describe fundamental relationships between protein structure, dynamics and function. Here we present an ensemble of ubiquitin in solution that is created by sampling conformational space without experimental information using "Backrub" motions inspired by alternative conformations observed in sub-Angstrom resolution crystal structures. Backrub-generated structures are then selected to produce an ensemble that optimizes agreement with nuclear magnetic resonance (NMR) Residual Dipolar Couplings (RDCs). Using this ensemble, we probe two proposed relationships between properties of protein ensembles: (i) a link between native-state dynamics and the conformational heterogeneity observed in crystal structures, and (ii) a relation between dynamics of an individual protein and the conformational variability explored by its natural family. We show that the Backrub motional mechanism can simultaneously explore protein native-state dynamics measured by RDCs, encompass the conformational variability present in ubiquitin complex structures and facilitate sampling of conformational and sequence variability matching those occurring in the ubiquitin protein family. Our results thus support an overall relation between protein dynamics and conformational changes enabling sequence changes in evolution. More practically, the presented method can be applied to improve protein design predictions by accounting for intrinsic native-state dynamics
Mining electron density for functionally relevant protein polysterism in crystal structures.
This review focuses on conceptual and methodological advances in our understanding and characterization of the conformational heterogeneity of proteins. Focusing on X-ray crystallography, we describe how polysterism, the interconversion of pre-existing conformational substates, has traditionally been analyzed by comparing independent crystal structures or multiple chains within a single crystal asymmetric unit. In contrast, recent studies have focused on mining electron density maps to reveal previously 'hidden' minor conformational substates. Functional tests of the importance of minor states suggest that evolutionary selection shapes the entire conformational landscape, including uniquely configured conformational substates, the relative distribution of these substates, and the speed at which the protein can interconvert between them. An increased focus on polysterism may shape the way protein structure and function is studied in the coming years
CABS-flex predictions of protein flexibility compared with NMR ensembles
Motivation: Identification of flexible regions of protein structures is
important for understanding of their biological functions. Recently, we have
developed a fast approach for predicting protein structure fluctuations from a
single protein model: the CABS-flex. CABS-flex was shown to be an efficient
alternative to conventional all-atom molecular dynamics (MD). In this work, we
evaluate CABS-flex and MD predictions by comparison with protein structural
variations within NMR ensembles.
Results: Based on a benchmark set of 140 proteins, we show that the relative
fluctuations of protein residues obtained from CABS-flex are well correlated to
those of NMR ensembles. On average, this correlation is stronger than that
between MD and NMR ensembles. In conclusion, CABS-flex is useful and
complementary to MD in predicting of protein regions that undergo
conformational changes and the extent of such changes
Electrostatic effects on funneled landscapes and structural diversity in denatured protein ensembles
The denatured state of proteins is heterogeneous and susceptible to general hydrophobic and electrostatic forces, but to what extent does the funneled nature of protein energy landscapes play a role in the unfolded ensemble? We simulate the denatured ensemble of cytochrome c using a series of models. The models pinpoint the efficacy of incorporating energetic funnels toward the native state in contrast with models having no native structure-seeking tendency. These models also contain varying strengths of electrostatic effects and hydrophobic collapse. The simulations based on these models are compared with experimental distributions for the distances between a fluorescent donor and the heme acceptor that were extracted from time-resolved fluorescence energy transfer experiments on cytochrome c. Comparing simulations to detailed experimental data on several labeling sites allows us to quantify the dominant forces in denatured protein ensembles
Structure and kinetics of chemically cross-linked protein gels from small-angle X-ray scattering
Glutaraldehyde (GA) reacts with amino groups in proteins, forming
intermolecular cross-links that, at sufficiently high protein concentration,
can transform a protein solution into a gel. Although GA has been used as a
cross-linking reagent for decades, neither the cross-linking chemistry nor the
microstructure of the resulting protein gel have been clearly established. Here
we use small-angle X-ray scattering (SAXS) to characterise the microstructure
and structural kinetics of gels formed by cross-linking of pancreatic trypsin
inhibitor, myoglobin or intestinal fatty acid-binding protein. By comparing the
scattering from gels and dilute solutions, we extract the structure factor and
the pair correlation function of the gels. The protein gels are spatially
heterogeneous, with dense clusters linked by sparse networks. Within the
clusters, adjacent protein molecules are almost in contact, but the protein
concentration in the cluster is much lower than in a crystal. At the 1 nm
SAXS resolution, the native protein structure is unaffected by cross-linking.
The cluster radius is in the range 10 - 50 nm, with the cluster size determined
mainly by the availability of lysine amino groups on the protein surface. The
development of structure in the gel, on time scales from minutes to hours,
appears to obey first-order kinetics. Cross-linking is slower at acidic pH,
where the population of amino groups in the reactive deprotonated form is low.
These results support the use of cross-linked protein gels in NMR studies of
protein dynamics and for modeling NMR relaxation in biological tissue.Comment: 16 pages, 11 figure
- …