The effects of telehealth running gait retraining on biomechanics, pain, and function in patients with lower extremity injuries: A randomized clinical trial.

Background: In-clinic gait retraining has been effective in modifying suspected biomechanical risk factors for running injury, but its feasibility is often limited by multiple clinic visits. This randomized clinical trial investigated the effects of a telehealth-based gait retraining intervention on running biomechanics, pain, and function in previously injured runners.

Methods: Twenty-three participants recovering from lower extremity injuries were randomized to a control or intervention group.

Principles of target DNA cleavage and the role of Mg2+ in the catalysis of CRISPR-Cas9.

At the core of the CRISPR-Cas9 genome-editing technology, the endonuclease Cas9 introduces site-specific breaks in DNA. However, precise mechanistic information to ameliorating Cas9 function is still missing. Here, multi-microsecond molecular dynamics, free-energy and multiscale simulations are combined with solution NMR and DNA cleavage experiments to resolve the catalytic mechanism of target DNA cleavage.

Identification and mechanistic basis of non-ACE2 blocking neutralizing antibodies from COVID-19 patients with deep RNA sequencing and molecular dynamics simulations.

Variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continue to cause disease and impair the effectiveness of treatments. The therapeutic potential of convergent neutralizing antibodies (NAbs) from fully recovered patients has been explored in several early stages of novel drugs. Here, we identified initially elicited NAbs (Ig Heavy, Ig lambda, Ig kappa) in response to COVID-19 infection in patients admitted to the intensive care unit at a single center with deep RNA sequencing (>100 million reads) of peripheral blood as a diagnostic tool for predicting the severity of the disease and as a means to pinpoint specific compensatory NAb treatments.

Enhanced specificity mutations perturb allosteric signaling in CRISPR-Cas9.

CRISPR-Cas9 (clustered regularly interspaced short palindromic repeat and associated Cas9 protein) is a molecular tool with transformative genome editing capabilities. At the molecular level, an intricate allosteric signaling is critical for DNA cleavage, but its role in the specificity enhancement of the Cas9 endonuclease is poorly understood. Here, multi-microsecond molecular dynamics is combined with solution NMR and graph theory-derived models to probe the allosteric role of key specificity-enhancing mutations.

A simple approach for reconstruction of non-uniformly sampled pseudo-3D NMR data for accurate measurement of spin relaxation parameters.

We explain how to conduct a pseudo-3D relaxation series NUS measurement so that it can be reconstructed by existing 3D NUS reconstruction methods to give accurate relaxation values. We demonstrate using reconstruction algorithms IST and SMILE that this 3D approach allows lower sampling densities than for independent 2D reconstructions. This is in keeping with the common finding that higher dimensionality increases signal sparsity, enabling lower sampling density.

H, C, N backbone resonance assignment of the recognition lobe subdomain 3 (Rec3) from Streptococcus pyogenes CRISPR-Cas9.

Rec3 is a subdomain of the recognition (Rec) lobe within CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-associated protein Cas9 that is involved in nucleic acid binding and is critical to HNH endonuclease activation. Here, we report the backbone resonance assignments of an engineered construct of the Rec3 subdomain from Streptococcus pyogenes Cas9. We also analyze backbone chemical shift data to predict secondary structure and an overall fold that is consistent with that of Rec3 from the full-length S.

Simultaneous Bilateral Fractures of the Tali.

A 22-year-old male US Military Academy cadet fell while sidestepping across the 8-ft-high bar portion of the indoor obstacle course. The cadet, in immense pain, was unable to bear weight immediately after the fall. Following examination by physical therapists within the fitness center, emergency medical service personnel transported the cadet to the emergency department for definitive care.

Acute Fracture of the Third Metacarpal in a Handball Player.

A 20-year-old right hand-dominant male military cadet presented to the direct-access physical therapy clinic complaining of pain and swelling of his right hand, which was injured while competing in a team handball match the day before. Due to suspicion of a third metacarpal fracture, fluoroscopy was performed on the cadet's hand in the physical therapy clinic, and an apparent oblique fracture was noted. Confirmatory radiographs were ordered and the cadet was referred for orthopaedic consultation.

Allosteric Motions of the CRISPR-Cas9 HNH Nuclease Probed by NMR and Molecular Dynamics.

CRISPR-Cas9 is a widely employed genome-editing tool with functionality reliant on the ability of the Cas9 endonuclease to introduce site-specific breaks in double-stranded DNA. In this system, an intriguing allosteric communication has been suggested to control its DNA cleavage activity through flexibility of the catalytic HNH domain. Here, solution NMR experiments and a novel Gaussian-accelerated molecular dynamics (GaMD) simulation method are used to capture the structural and dynamic determinants of allosteric signaling within the HNH domain.

NMR and computational methods for molecular resolution of allosteric pathways in enzyme complexes.

Allostery is a ubiquitous biological mechanism in which a distant binding site is coupled to and drastically alters the function of a catalytic site in a protein. Allostery provides a high level of spatial and temporal control of the integrity and activity of biomolecular assembles composed of proteins, nucleic acids, or small molecules. Understanding the physical forces that drive allosteric coupling is critical to harnessing this process for use in bioengineering, de novo protein design, and drug discovery.

H, C, N backbone and side chain resonance assignment of the HNH nuclease from Streptococcus pyogenes CRISPR-Cas9.

HNH is one of two endonuclease domains of the clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein Cas9 that perform site-specific cleavage of double-stranded DNA. We engineered a novel construct of this critical nuclease from Streptococcus pyogenes Cas9 that not only maintains the wild-type amino acid sequence and fold, but displays enhanced thermostability when compared to the full-length Cas9 enzyme. Here, we report backbone and side chain assignments of the HNH nuclease as a foundational step toward the characterization of protein dynamics and allostery in CRISPR-Cas9.

I260Q DNA polymerase β highlights precatalytic conformational rearrangements critical for fidelity.

DNA polymerase β (pol β) fills single nucleotide gaps in DNA during base excision repair and non-homologous end-joining. Pol β must select the correct nucleotide from among a pool of four nucleotides with similar structures and properties in order to maintain genomic stability during DNA repair. Here, we use a combination of X-ray crystallography, fluorescence resonance energy transfer and nuclear magnetic resonance to show that pol β's ability to access the appropriate conformations both before and upon binding to nucleotide substrates is integral to its fidelity.

Altering the allosteric pathway in IGPS suppresses millisecond motions and catalytic activity.

Imidazole glycerol phosphate synthase (IGPS) is a V-type allosteric enzyme, meaning that its catalytic rate is critically dependent on activation by its allosteric ligand, -[(5'-phosphoribulosyl)formimino]-5-aminoimidazole-4-carboxamide ribonucleotide (PRFAR). The allosteric mechanism of IGPS is reliant on millisecond conformational motions for efficient catalysis. We engineered four mutants of IGPS designed to disrupt millisecond motions and allosteric coupling to identify regions that are critical to IGPS function.

Brief early-life non-specific incorporation of deuterium extends mean life span in Drosophila melanogaster without affecting fecundity.

We have investigated the effects of brief, non-specific deuteration of Drosophila melanogaster by including varying percentages of ²H (D) in the H₂O used in the food mix consumed during initial development. Up to 22.5% deuterium oxide (D₂O) in H₂O was administered, with the result that a low percentage of D₂O in the water increased mean life span, whereas the highest percentage used (22.