Cooperative protein allosteric transition mediated by a fluctuating transmission network

Allosteric communication between distant protein sites represents a key mechanism of biomolecular regulation and signal transduction. Compared to other processes such as protein folding, however, the dynamical evolution of allosteric transitions is still not well understood. As example of allosteric coupling between distant protein regions, we consider the global open-closed motion of the two domains of T4 lysozyme, which is triggered by local motions in the hinge region. Combining extensive molecular dynamics simulations with machine learning of contact features, we identify a network of interresidue distances that move in a concerted manner. The cooperative process originates from a cogwheel-like motion of the hydrophobic core in the hinge region, which constitutes a flexible transmission network. Through rigid contacts and the protein backbone, the small local changes of the hydrophobic core are passed on to the distant terminal domains and lead to the emergence of a rare global conformational transition. As in an Ising-type model, the cooperativity of the allosteric transition can be explained via the interaction of local fluctuations.

First Crystal Structure of Bacterial Oligopeptidase B in an Intermediate State: The Roles of the Hinge Region Modification and Spermine

Oligopeptidase B (OpB) is a two-domain, trypsin-like serine peptidase belonging to the S9 prolyloligopeptidase (POP) family. Two domains are linked by a hinge region that participates in the transition of the enzyme between two major states—closed and open—in which domains and residues of the catalytic triad are located close to each other and separated, respectively. In this study, we described, for the first time, a structure of OpB from bacteria obtained for an enzyme from Serratia proteomaculans with a modified hinge region (PSPmod). PSPmod was crystallized in a conformation characterized by a disruption of the catalytic triad together with a domain arrangement intermediate between open and closed states found in crystals of ligand-free and inhibitor-bound POP, respectively. Two additional derivatives of PSPmod were crystallized in the same conformation. Neither wild-type PSP nor its corresponding mutated variants were susceptible to crystallization, indicating that the hinge region modification was key in the crystallization process. The second key factor was suggested to be polyamine spermine since all crystals were grown in its presence. The influences of the hinge region modification and spermine on the conformational state of PSP in solution were evaluated by small-angle X-ray scattering. SAXS showed that, in solution, wild-type PSP adopted the open state, spermine caused the conformational transition to the intermediate state, and spermine-free PSPmod contained molecules in the open and intermediate conformations in dynamic equilibrium.

Seismic Behavior of RC Bridge Piers Locally Replaced with SFRC-FA Subjected to Torsion Combined with Axial Compression

Under complex seismic forces, the failure characteristics of the plastic hinge region at the bottom of the pier column and the methods improving the ductility have attracted extensive attention. In this study, steel fiber-reinforced concrete with fine aggregate (SFRC-FA) was applied to locally replace the conventional concrete in the potential plastic hinge region at the bottom of the pier column. Five SFRC-FA pier column specimens with different stirrup ratios and different replacement lengths and one conventional reinforced concrete pier column specimen were produced. Using the seismic behavior tests under the combined bending-shear-torsion-axial force, the failure mode, torsional bearing capacity, energy dissipation, and the torsional plastic hinges of the pier columns were investigated. In addition, an equation for calculating the torsional bearing capacity of the new composite pier columns was proposed. The results showed that (1) compared with the reinforced concrete pier column, the plastic hinge was shifted from the bottom of the pier column to the middle of the height of the pier column due to the application of SFRC-FA at the bottom of the pier column, which improved the torsional bearing capacity; (2) the effect of reducing the stirrup ratio of the SFRC-FA replacement region on the torsional bearing capacity, cracking mode, energy dissipation, and ductility was not obvious; (3) the accuracy of the new equation based on the space truss model proposed in this article was verified by comparison with the experiments of this study and other researches.

Cinnamamide-Chalcone Derivatives as CDK2 Inhibitors: Synthesis, Pharmacological Evaluation, and Molecular Modelling Study

A series of 13 novel cinnamamide-chalcone derivatives (2a-2m) were synthesized and evaluated for their antiproliferative activity against MCF-7, K562, U373MG, and HT-29 cell lines by SRB assay. Considering the activities on MCF-7 cell line, eight compounds were tested for the in-vitro CDK2 inhibition and four (2g, 2h, 2k and 2l) were found to possess good activity (IC50<10µM). These four compounds were tested on EGFR kinase to assess the selectivity towards CDK2 and were found be nearly two times more selective. To corroborate the in-vitro enzyme assay data with binding, the compounds were docked into the CDK2 and EGFR using Glide software. The docking studies reveal that all eight compounds form hydrogen bonds with Lys33 (β-3 region) and Leu83 (hinge region) in CDK2 and the docking scores correlate well with the IC50 values. The most active compounds on CDK2 when docked in EGFR had lower docking scores. Only one compound interacts with Lys721 (β-3 region) and Met769 (hinge region). The stability of interactions with CDK2 was assessed for 2k and 2l by molecular dynamics simulation using Desmond software. In conclusion, three compounds possess excellent activity against MCF-7 cell line and good activity against CDK2.

Classification, structural biology, and applications of mucin domain-targeting proteases

Epithelial surfaces throughout the body are coated by mucins, a class of proteins carrying domains characterized by a high density of O-glycosylated serine and threonine residues. The resulting mucosal layers form crucial host-microbe interfaces that prevent the translocation of microbes while also selecting for distinct bacteria via the presented glycan repertoire. The intricate interplay between mucus production and breakdown thus determines the composition of the microbiota maintained within these mucosal environments, which can have a large influence on the host during both homeostasis and disease. Most research to date on mucus breakdown has focused on glycosidases that trim glycan structures to release monosaccharides as a source of nutrients. More recent work has uncovered the existence of mucin-type O-glycosylation-dependent proteases that are secreted by pathogens, commensals, and mutualists to facilitate mucosal colonization and penetration. Additionally, immunoglobulin A (IgA) proteases promote bacterial colonization in the presence of neutralizing secretory IgA through selective cleavage of the heavily O-glycosylated hinge region. In this review, we summarize families of O-glycoproteases and IgA proteases, discuss known structural features, and review applications of these enzymes to glycobiology.

D614G substitution at the hinge region enhances the stability of trimeric SARS-CoV-2 spike protein

Mutations in the spike protein of SARS-CoV-2 are the major causes for the modulation of ongoing COVID-19 infection. Currently, the D614G substitution in the spike protein has become dominant worldwide. It is associated with higher infectivity than the ancestral (D614) variant. We demonstrate using Gaussian network model-based normal mode analysis that the D614G substitution occurs at the hinge region that facilitates domain-domain motions between receptor binding domain and S2 region of the spike protein. Computer-aided mutagenesis and inter-residue energy calculations reveal that contacts involving D614 are energetically frustrated. However, contacts involving G614 are energetically favourable, implying the substitution strengthens residue contacts that are formed within as well as between protomers. We also find that the free energy difference (ΔΔG) between two variants is -2.6 kcal/mol for closed and -2.0 kcal/mol for 1-RBD up conformation. Thus, the hermodynamic stability has increased upon D614G substitution. Whereas the reverse mutation in spike protein structures having G614 substitution has resulted in the free energy differences of 6.6 kcal/mol and 6.3 kcal/mol for closed and 1-RBD up conformations, respectively, indicating that the overall thermodynamic stability has decreased. These results suggest that the D614G substitution modulates the flexibility of spike protein and confers enhanced thermodynamic stability irrespective of conformational states. This data concurs with the known information demonstrating increased availability of the functional form of spike protein trimer upon D614G substitution.