OpenTn5: Open-Source Resource for Robust and Scalable Tn5 Transposase Purification and Characterization.

Tagmentation combines DNA fragmentation and sequencing adapter addition by leveraging the transposition activity of the bacterial cut-and-paste Tn5 transposase, to enable efficient sequencing library preparation. Here we present an open-source protocol for the generation of multi-purpose hyperactive Tn5 transposase, including its benchmarking in CUT&Tag, bulk and single-cell ATAC-seq. The OpenTn5 protocol yields multi-milligram quantities of pG-Tn5 protein per liter of culture, sufficient for thousands of tagmentation reactions and the enzyme retains activity in storage for more than a year.

Mechanism of glycoform specificity and in vivo protection by an anti-afucosylated IgG nanobody.

Immunoglobulin G (IgG) antibodies contain a complex N-glycan embedded in the hydrophobic pocket between its heavy chain protomers. This glycan contributes to the structural organization of the Fc domain and determines its specificity for Fcγ receptors, thereby dictating distinct cellular responses. The variable construction of this glycan structure leads to highly-related, but non-equivalent glycoproteins known as glycoforms.

Itch receptor MRGPRX4 interacts with the receptor activity-modifying proteins.

Cholestatic itch is a severe and debilitating symptom in liver diseases with limited treatment options. The class A G protein-coupled receptor (GPCR) Mas-related GPCR subtype X4 (MRGPRX4) has been identified as a receptor for bile acids, which are potential cholestatic pruritogens. An increasing number of GPCRs have been shown to interact with receptor activity-modifying proteins (RAMPs), which can modulate different aspects of GPCR biology.

Mechanism of glycoform specificity and protection against antibody dependent enhancement by an anti-afucosylated IgG nanobody.

Immunoglobulin G (IgG) antibodies contain a single, complex -glycan on each IgG heavy chain protomer embedded in the hydrophobic pocket between its Cγ2 domains. The presence of this glycan contributes to the structural organization of the Fc domain and determines its specificity for Fcγ receptors, thereby determining distinct cellular responses. On the Fc, the variable construction of this glycan structure leads to a family of highly-related, but non-equivalent glycoproteins known as glycoforms.

Enhanced SARS-CoV-2 neutralization by dimeric IgA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus that causes coronavirus disease 2019 (COVID-19), primarily infects cells at mucosal surfaces. Serum neutralizing antibody responses are variable and generally low in individuals that suffer mild forms of COVID-19. Although potent immunoglobulin G (IgG) antibodies can neutralize the virus, less is known about secretory antibodies such as IgA that might affect the initial viral spread and transmissibility from the mucosa.

Enhanced SARS-CoV-2 Neutralization by Secretory IgA in vitro.

SARS-CoV-2 primarily infects cells at mucosal surfaces. Serum neutralizing antibody responses are variable and generally low in individuals that suffer mild forms of the illness. Although potent IgG antibodies can neutralize the virus, less is known about secretory antibodies such as IgA that might impact the initial viral spread and transmissibility from the mucosa.

A Combination of Human Broadly Neutralizing Antibodies against Hepatitis B Virus HBsAg with Distinct Epitopes Suppresses Escape Mutations.

Although there is no effective cure for chronic hepatitis B virus (HBV) infection, antibodies are protective and correlate with recovery from infection. To examine the human antibody response to HBV, we screened 124 vaccinated and 20 infected, spontaneously recovered individuals. The selected individuals produced shared clones of broadly neutralizing antibodies (bNAbs) that targeted 3 non-overlapping epitopes on the HBV S antigen (HBsAg).

Carbon monoxide: a critical physiological regulator sensitive to light.

The mechanism by which humans absorb therapeutic light in winter seasonal and nonseasonal depression is unknown. Bright-light-induced release and generation of blood-borne gasotransmitters such as carbon monoxide (CO) may be one mechanism. Here, 24 healthy female volunteers had peripheral blood samples drawn.

secreted antigen A generates muropeptides to enhance host immunity and limit bacterial pathogenesis.

We discovered that (), a ubiquitous commensal bacterium, and its secreted peptidoglycan hydrolase (SagA) were sufficient to enhance intestinal barrier function and pathogen tolerance, but the precise biochemical mechanism was unknown. Here we show has unique peptidoglycan composition and remodeling activity through SagA, which generates smaller muropeptides that more effectively activates nucleotide-binding oligomerization domain-containing protein 2 (NOD2) in mammalian cells. Our structural and biochemical studies show that SagA is a NlpC/p60-endopeptidase that preferentially hydrolyzes crosslinked Lys-type peptidoglycan fragments.

Identification of a Mycothiol-Dependent Nitroreductase from Mycobacterium tuberculosis.

The success of Mycobacterium tuberculosis (Mtb) as a pathogen depends on the redundant and complex mechanisms it has evolved for resisting nitrosative and oxidative stresses inflicted by host immunity. Improving our understanding of these defense pathways can reveal vulnerable points in Mtb pathogenesis. In this study, we combined genetic, structural, computational, biochemical, and biophysical approaches to identify a novel enzyme class represented by Rv2466c.

Retinal venous blood carbon monoxide response to bright light in male pigs: A preliminary study.

The physical mechanism by which light is absorbed in the eye and has antidepressant and energizing effects in Seasonal Affective Disorder and other forms of psychiatric major depression is of scientific interest. This study was designed to explore one specific aspect of a proposed humoral phototransduction mechanism, namely that carbon monoxide (CO) levels increase in retinal venous blood in response to bright light. Eleven mature male pigs approximately six months of age were kept for 7days in darkness and fasted for 12h prior to surgery.

Structural Mapping and Functional Characterization of Zebrafish Class B G-Protein Coupled Receptor (GPCR) with Dual Ligand Selectivity towards GLP-1 and Glucagon.

GLP-1 and glucagon regulate glucose metabolism through a network of metabolic pathways initiated upon binding to their specific receptors that belong to class B G-protein coupled receptors (GPCRs). The therapeutic potential of glucagon is currently being evaluated, while GLP-1 is already used in the treatment of type 2 diabetes and obesity. Development of a second generation of GLP-1 based therapeutics depends on a molecular and structural understanding of the interactions between the GLP-1 receptor (GLP-1R) and its ligand GLP-1.

Biochemical and structural analysis of the interaction between β-amyloid and fibrinogen.

The majority of patients with Alzheimer disease (AD) suffer from impaired cerebral circulation. Accumulating evidence suggests that fibrinogen, the main protein component of blood clots, plays an important role in this circulatory dysfunction in AD. Fibrinogen interacts with β-amyloid (Aβ), forming plasmin-resistant abnormal blood clots, and increased fibrin deposition is found in the brains of AD patients and mouse models.

Changes in melatonin synthesis parameters after carbon monoxide concentration increase in the cavernous sinus.

Previous studies indicate that the gaseous messenger carbon monoxide (CO) is released from the eye into the ophthalmic venous blood depending on the intensity of sunlight. This study was designed to determine whether the increased concentration of CO in ophthalmic venous blood affects the synthesis of melatonin and therefore, whether CO released from the eye under normal lighting conditions can be a carrier of light intensity information. Thirty six mature male wild boar and pig crossbreeds (n = 36) were studied.

Strategic addition of an N-linked glycan to a monoclonal antibody improves its HIV-1-neutralizing activity.

Ibalizumab is a humanized monoclonal antibody that binds human CD4--a key receptor for HIV--and blocks HIV-1 infection. However, HIV-1 strains with mutations resulting in loss of an N-linked glycan from the V5 loop of the envelope glycoprotein gp120 are resistant to ibalizumab. Previous structural analysis suggests that this glycan fills a void between the gp120 V5 loop and the ibalizumab light chain, perhaps causing steric hindrance that disrupts viral entry.

SAD and the not-so-single photoreceptors.

Research in the last century has demonstrated that light is a critical regulator of physiology in animals. More recent research has exposed the influence of light on human behavior, including the phenomenon of seasonal affective disorder (SAD). Repeated studies have shown that light treatment is effective in this disorder.

Bispecific antibodies directed to CD4 domain 2 and HIV envelope exhibit exceptional breadth and picomolar potency against HIV-1.

In the absence of an effective HIV-1 vaccine, passive immunization using broadly neutralizing Abs or Ab-like molecules could provide an alternative to the daily administration of oral antiretroviral agents that has recently shown promise as preexposure prophylaxis. Currently, no single broadly neutralizing Ab (bNAb) or combination of bNAbs neutralizes all HIV-1 strains at practically achievable concentrations in vivo. To address this problem, we created bispecific Abs that combine the HIV-1 inhibitory activity of ibalizumab (iMab), a humanized mAb directed to domain 2 of human CD4, with that of anti-gp120 bNAbs.

From screen to structure with a harvestable microfluidic device.

Advances in automation have facilitated the widespread adoption of high-throughput vapour-diffusion methods for initial crystallization screening. However, for many proteins, screening thousands of crystallization conditions fails to yield crystals of sufficient quality for structural characterization. Here, the rates of crystal identification for thaumatin, catalase and myoglobin using microfluidic Crystal Former devices and sitting-drop vapour-diffusion plates are compared.

Multicenter, randomized, double-blind, active comparator and placebo-controlled trial of a corticotropin-releasing factor receptor-1 antagonist in generalized anxiety disorder.

Background: Antagonism of corticotropin-releasing factor (CRF) receptors has been hypothesized as a potential target for the development of novel anxiolytics. This study was designed to determine the safety and efficacy of pexacerfont, a selective CRF-1 receptor antagonist, in the treatment of generalized anxiety disorder (GAD).

Method: This was a multicenter, randomized, double-blind, placebo-controlled and active comparator trial.

Aripiprazole monotherapy in acute mania: 12-week randomised placebo- and haloperidol-controlled study.

Background: Well-tolerated and effective therapies for bipolar mania are required.

Aims: To evaluate the efficacy and tolerability of aripiprazole as acute and maintenance of effect therapy in patients with bipolar I disorder experiencing manic or mixed episodes.

Method: Patients were randomised to double-blind aripiprazole (15 or 30 mg/day; n=167), placebo (n=153) or haloperidol (5-15 mg/day, n=165) for 3 weeks (trial registration NCT00097266).

A randomized, double-blind, placebo-controlled tolerability study of intramuscular aripiprazole in acutely agitated patients with Alzheimer's, vascular, or mixed dementia.

Objectives: To evaluate the tolerability of intramuscular (IM) aripiprazole in patients with agitation associated with dementia.

Design: A 24-hour, double-blind, placebo-controlled, randomized study.

Setting: Sixteen healthcare facilities in the United States.

Crystal engineering of HIV-1 reverse transcriptase for structure-based drug design.

HIV-1 reverse transcriptase (RT) is a primary target for anti-AIDS drugs. Structures of HIV-1 RT, usually determined at approximately 2.5-3.

Treatment effects of selegiline transdermal system on symptoms of major depressive disorder: a meta-analysis of short-term, placebo-controlled, efficacy trials.

Objective: Selegiline transdermal system (STS) is efficacious for the treatment of major depressive disorder (MDD). This meta-analysis explores treatment effects of STS for individual symptoms of MDD derived from line-item analyses of the 28-item Hamilton Rating Scale for Depression (HAM-D28) and the Montgomery-Asberg Depression Rating Scale (MADRS).

Methods: Change in score from baseline to end of treatment for each item of the HAM-D28 and MADRS was assessed using a multilevel model for meta-analysis of continuous outcome data from all five short-term, randomized, placebo-controlled efficacy trials conducted during preapproval clinical development of STS for MDD.

Management of acute agitation in patients with bipolar disorder: efficacy and safety of intramuscular aripiprazole.

To investigate the efficacy and safety of intramuscular (IM) aripiprazole for the treatment of agitation in patients with bipolar I disorder, manic or mixed episodes. In total, 301 patients experiencing acute agitation were randomized to IM aripiprazole 9.75 mg per injection (n = 78), IM aripiprazole 15 mg per injection (n = 78), IM lorazepam 2 mg per injection (n = 70), or IM placebo (n = 75) in this double-blind multicenter study.