Small Peptide Binding Stiffens the Ubiquitin-like Protein SUMO1

AbstractPosttranslational modification by small ubiquitin-like modifiers (SUMOs), known as SUMOylation, is a key regulatory event in many eukaryotic cellular processes in which SUMOs interact with a large number of target proteins. SUMO binding motifs (SBMs) are small peptides derived from these target proteins that interact noncovalently with SUMOs and induce conformational changes. To determine the effect of SBMs on the mechanical properties of SUMO1 (the first member of the human SUMO family), we performed single-molecule force spectroscopy experiments on SUMO1/SBM complexes. The unfolding force of SUMO1 (at a pulling speed of 400 nm/s) increased from ∼130 pN to ∼170 pN upon binding to SBMs, indicating mechanical stabilization upon complexation. Pulling-speed-dependent experiments and Monte Carlo simulations measured a large decrease in distance to the unfolding transition state for SUMO1 upon SBM binding, which is by far the largest change measured for any ligand binding protein. The stiffness of SUMO1 (measured as a spring constant for the deformation response along the line joining the N- and C-termini) increased upon SBM binding from ∼1 N/m to ∼3.5 N/m. The relatively higher flexibility of ligand-free SUMO1 might play a role in accessing various conformations before binding to a target

Role of conservative mutations in protein multi-property adaptation

Protein physicochemical properties must undergo complex changes during evolution, as a response to modifications in the organism environment, the result of the proteins taking up new roles or because of the need to cope with the evolution of molecular interacting partners. Recent work has emphasized the role of stability and stability–function trade-offs in these protein adaptation processes. In the present study, on the other hand, we report that combinations of a few conservative, high-frequency-of-fixation mutations in the thioredoxin molecule lead to largely independent changes in both stability and the diversity of catalytic mechanisms, as revealed by single-molecule atomic force spectroscopy. Furthermore, the changes found are evolutionarily significant, as they combine typically hyperthermophilic stability enhancements with modulations in function that span the ranges defined by the quite different catalytic patterns of thioredoxins from bacterial and eukaryotic origin. These results suggest that evolutionary protein adaptation may use, in some cases at least, the potential of conservative mutations to originate a multiplicity of evolutionarily allowed mutational paths leading to a variety of protein modulation patterns. In addition the results support the feasibility of using evolutionary information to achieve protein multi-feature optimization, an important biotechnological goal

Family Firms and Firm Performance: Evidence from Japan

Corrigendum: Nature Structural and Molecular Biology 16 (12), 1331 (2009) doi:10.1038/nsmb1209-1331bInternational audienceThioredoxins (Trxs) are oxidoreductase enzymes, present in all organisms, that catalyze the reduction of disulfide bonds in proteins. By applying a calibrated force to a substrate disulfide, the chemical mechanisms of Trx catalysis can be examined in detail at the single-molecule level. Here we use single-molecule force-clamp spectroscopy to explore the chemical evolution of Trx catalysis by probing the chemistry of eight different Trx enzymes. All Trxs show a characteristic Michaelis-Menten mechanism that is detected when the disulfide bond is stretched at low forces, but at high forces, two different chemical behaviors distinguish bacterial-origin from eukaryotic-origin Trxs. Eukaryotic-origin Trxs reduce disulfide bonds through a single-electron transfer reaction (SET), whereas bacterial-origin Trxs show both nucleophilic substitution (SN2) and SET reactions. A computational analysis of Trx structures identifies the evolution of the binding groove as an important factor controlling the chemistry of Trx catalysis

Variance of Atomic Coordinates as a Dynamical Metric to Distinguish Proteins and Protein-Protein Interactions in Molecular Dynamics Simulations

In this manuscript, we introduce the Cumulative Variance of Coordinate Fluctuations (CVCF) along atomistic MD trajectories, as a dynamical metric to examine protein dynamics and sampling convergence in MD simulations. Using model 1D and 2D PES, we first show that CVCF, which traces over the fluctuations of protein atoms as a function of sampling coordinate (time in MD simulations), captures both local and global equilibria to distinguish the underlying PES of proteins. For both model PES and protein trajectories, we compare the information content present in CVCF traces with that obtained using other measures proposed in literature to reveal conditions under which a consistent interpretation of data can be obtained. Importantly, we show that independent of convergence to either local or global equilibrium, the values and features of protein CVCF can provide a comparative assessment of the ruggedness and curvature of the underlying PES sampled by proteins along MD trajectories. Trends in CVCF therefore enable us to compare features of the PES across multiple protein systems using MD simulations. We demonstrate some of the attractive features of a CVCF based analysis on multi-microsecond (ms) MD trajectories of structurally homologous ubiquitin family proteins which present a particularly striking example in nature wherein sequence changes and complexation which do not lead to prominent structural changes bring about dramatic functional consequences