Structural and functional characterization of the nucleotide-binding domains of ABCA4 and their role in Stargardt disease.

ABCA4 is an ATP-binding cassette (ABC) transporter that prevents the buildup of toxic retinoid compounds by facilitating the transport of N-retinylidene-phosphatidylethanolamine across membranes of rod and cone photoreceptor cells. Over 1500 missense mutations in ABCA4, many in the nucleotide-binding domains (NBDs), have been genetically linked to Stargardt disease. Here, we show by cryo-EM that ABCA4 is converted from an open outward conformation to a closed conformation upon the binding of adenylyl-imidodiphosphate.

Next generation of multispecific antibody engineering.

Multispecific antibodies recognize two or more epitopes located on the same or distinct targets. This added capability through protein design allows these man-made molecules to address unmet medical needs that are no longer possible with single targeting such as with monoclonal antibodies or cytokines alone. However, the approach to the development of these multispecific molecules has been met with numerous road bumps, which suggests that a new workflow for multispecific molecules is required.

Beyond S. cerevisiae for winemaking: Fermentation-related trait diversity in the genus Saccharomyces.

Saccharomyces cerevisiae is the yeast of choice for most inoculated wine fermentations worldwide. However, many other yeast species and genera display phenotypes of interest that may help address the environmental and commercial challenges the wine industry has been facing in recent years. This work aimed to provide, for the first time, a systematic phenotyping of all Saccharomyces species under winemaking conditions.

Protocol for high-throughput cloning, expression, purification, and evaluation of bispecific antibodies.

Bispecific antibodies are a powerful new class of therapeutics, but their development often requires enormous amounts of time and resources. Here, we describe a high-throughput protocol for cloning, expressing, purifying, and evaluating bispecific antibodies. This protocol enables the rapid screening of large panels of bispecific molecules to identify top candidates for further development.

Podoplanin Expression Independently and Jointly with Oral Epithelial Dysplasia Grade Acts as a Potential Biomarker of Malignant Transformation in Oral Leukoplakia.

Our aim was to evaluate the expression of biomarkers, CD44v6, CD147, EGFR, p53, p63, p73, p16, and podoplanin in oral leukoplakias (OL) and to assess their potential for prediction of malignant transformation (MT). We analyzed the expression of CD44v6, CD147, EGFR, p53, p63, p73, p16, and podoplanin by immunohistochemistry in 52 OL, comprised of 41 low-grade (LG) dysplasia and 11 high-grade (HG) cases. Twelve healthy normal tissues (NT) were also included.

LY-CoV1404 (bebtelovimab) potently neutralizes SARS-CoV-2 variants.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-neutralizing monoclonal antibodies (mAbs) can reduce the risk of hospitalization from coronavirus disease 2019 (COVID-19) when administered early. However, SARS-CoV-2 variants of concern (VOCs) have negatively affected therapeutic use of some authorized mAbs. Using a high-throughput B cell screening pipeline, we isolated LY-CoV1404 (bebtelovimab), a highly potent SARS-CoV-2 spike glycoprotein receptor binding domain (RBD)-specific antibody.

Structure and function of ABCA4 and its role in the visual cycle and Stargardt macular degeneration.

ABCA4 is a member of the superfamily of ATP-binding cassette (ABC) transporters that is preferentially localized along the rim region of rod and cone photoreceptor outer segment disc membranes. It uses the energy from ATP binding and hydrolysis to transport N-retinylidene-phosphatidylethanolamine (N-Ret-PE), the Schiff base adduct of retinal and phosphatidylethanolamine, from the lumen to the cytoplasmic leaflet of disc membranes. This ensures that all-trans-retinal and excess 11-cis-retinal are efficiently cleared from photoreceptor cells thereby preventing the accumulation of toxic retinoid compounds.

Next generation Fc scaffold for multispecific antibodies.

Bispecific antibodies (Bispecifics) demonstrate exceptional clinical potential to address some of the most complex diseases. However, Bispecific production in a single cell often requires the correct pairing of multiple polypeptide chains for desired assembly. This is a considerable hurdle that hinders the development of many immunoglobulin G (IgG)-like bispecific formats.

Expression of spindle assembly checkpoint proteins BubR1 and Mad2 expression as potential biomarkers of malignant transformation of oral leukoplakia: an observational cohort study.

Background: The Spindle Assembly Checkpoint (SAC) is a surveillance mechanism essential to ensure the accuracy of chromosome segregation during mitosis. Our aim was to evaluate the expression of SAC proteins in oral carcinogenesis, and to assess their potential in predicting malignant transformation of oral leukoplakia.

Material And Methods: We analysed the immunoexpression of BubR1, Mad2, Bub3, and Spindly proteins in 64 oral biopsies from 52 oral leukoplakias and 12 normal tissues.

Cryo-EM structures of the ABCA4 importer reveal mechanisms underlying substrate binding and Stargardt disease.

ABCA4 is an ATP-binding cassette (ABC) transporter that flips N-retinylidene-phosphatidylethanolamine (N-Ret-PE) from the lumen to the cytoplasmic leaflet of photoreceptor membranes. Loss-of-function mutations cause Stargardt disease (STGD1), a macular dystrophy associated with severe vision loss. To define the mechanisms underlying substrate binding and STGD1, we determine the cryo-EM structure of ABCA4 in its substrate-free and bound states.

LY-CoV1404 (bebtelovimab) potently neutralizes SARS-CoV-2 variants.

Unlabelled: SARS-CoV-2 neutralizing monoclonal antibodies (mAbs) can reduce the risk of hospitalization when administered early during COVID-19 disease. However, the emergence of variants of concern has negatively impacted the therapeutic use of some authorized mAbs. Using a high throughput B-cell screening pipeline, we isolated a highly potent SARS-CoV-2 spike glycoprotein receptor binding domain (RBD)-specific antibody called LY-CoV1404 (also known as bebtelovimab).

Enhancing the Prefusion Conformational Stability of SARS-CoV-2 Spike Protein Through Structure-Guided Design.

The worldwide pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is unprecedented and the impact on public health and the global economy continues to be devastating. Although early therapies such as prophylactic antibodies and vaccines show great promise, there are concerns about the long-term efficacy and universal applicability of these therapies as the virus continues to mutate. Thus, protein-based immunogens that can quickly respond to viral changes remain of continued interest.

Identification of critical chemical modifications and paratope mapping by size exclusion chromatography of stressed antibody-target complexes.

Therapeutic proteins including antibodies and Fc-fusion proteins undergo a large number of chemical modifications during cell culture, purification, storage and in human circulation. They are also exposed to harsh conditions during stress studies, including elevated temperature, extremes of pH, forced oxidation, physiological pH, UV light to assess the possible degradation pathways and suitability of methods for detecting them. Some of these modifications are located on residues in binding regions, leading to loss of binding and potency and classified as critical quality attributes.

Rational selection of building blocks for the assembly of bispecific antibodies.

Bispecific antibodies, engineered to recognize two targets simultaneously, demonstrate exceptional clinical potential for the therapeutic intervention of complex diseases. However, these molecules are often composed of multiple polypeptide chains of differing sequences. To meet industrial scale productivity, enforcing the correct quaternary assembly of these chains is critical.

Functional Characterization of ABCA4 Missense Variants Linked to Stargardt Macular Degeneration.

ABCA4 is an ATP-binding cassette (ABC) transporter expressed in photoreceptors, where it transports its substrate, -retinylidene-phosphatidylethanolamine (-Ret-PE), across outer segment membranes to facilitate the clearance of retinal from photoreceptors. Mutations in cause Stargardt macular degeneration (STGD1), an autosomal recessive disorder characterized by a loss of central vision and the accumulation of bisretinoid compounds. The purpose of this study was to determine the molecular properties of ABCA4 variants harboring disease-causing missense mutations in the transmembrane domains.

Functional analysis and classification of homozygous and hypomorphic ABCA4 variants associated with Stargardt macular degeneration.

Stargardt macular degeneration (Stargardt disease 1 [STGD1]) is caused by mutations in the gene encoding ABCA4, an ATP-binding cassette protein that transports N-retinylidene-phosphatidylethanolamine (N-Ret-PE) across photoreceptor membranes. Reduced ABCA4 activity results in retinoid accumulation leading to photoreceptor degeneration. The disease onset and severity vary from severe loss in visual acuity in the first decade to mild visual impairment late in life.

Molecular Insight into Recognition of the CGRPR Complex by Migraine Prevention Therapy Aimovig (Erenumab).

Calcitonin-gene-related peptide (CGRP) plays a key role in migraine pathophysiology. Aimovig (erenumab; erenumab-aooe in the United States) is the only US Food and Drug Administration (FDA)-approved monoclonal antibody (mAb) therapy against the CGRP receptor (CGRPR) for the prevention of migraine. Aimovig is also the first FDA-approved mAb against a G-protein-coupled receptor (GPCR).

The Chimpanzee SIV Envelope Trimer: Structure and Deployment as an HIV Vaccine Template.

Epitope-targeted HIV vaccine design seeks to focus antibody responses to broadly neutralizing antibody (bnAb) sites by sequential immunization. A chimpanzee simian immunodeficiency virus (SIV) envelope (Env) shares a single bnAb site, the variable loop 2 (V2)-apex, with HIV, suggesting its possible utility in an HIV immunization strategy. Here, we generate a chimpanzee SIV Env trimer, MT145K, which displays selective binding to HIV V2-apex bnAbs and precursor versions, but no binding to other HIV specificities.

A histological evaluation of the surgical margins from human oral fibrous-epithelial lesions excised with CO2 laser, Diode laser, Er:YAG laser, Nd:YAG laser, electrosurgical scalpel and cold scalpel.

Background: We aim to evaluate the presence of histological artefacts in the surgical margins of human oral fibro-epithelial hyperplasias excised with lasers of different wavelengths, and also electrosurgical scalpel and cold scalpel. Moreover, we aim to determine if some of these instruments could impair the normal histological diagnosis of these lesions.

Material And Methods: We included 130 consecutive surgical samples of 80 females and 50 males (mean age of 53.

[Informed Consent - Vision and Perspectives of Adolescents, Parents and Professionals: Multicentric Study in Six Hospitals].

Introduction: According to the Convention on the Rights of Children and the national standards of the Portuguese Directorate-General for Health, adolescents have the right to make decisions about their own health. The aim of this study was to identify the dynamics of the implementation of assent and informed consent in hospital settings.

Material And Methods: Cross-sectional and multicentre study based on surveys, which included adolescents from 14 to 18 years and their parents.

Efficient Single-Strand Break Repair Requires Binding to Both Poly(ADP-Ribose) and DNA by the Central BRCT Domain of XRCC1.

XRCC1 accelerates repair of DNA single-strand breaks by acting as a scaffold protein for the recruitment of Polβ, LigIIIα, and end-processing factors, such as PNKP and APTX. XRCC1 itself is recruited to DNA damage through interaction of its central BRCT domain with poly(ADP-ribose) chains generated by PARP1 or PARP2. XRCC1 is believed to interact directly with DNA at sites of damage, but the molecular basis for this interaction within XRCC1 remains unclear.

Correlating the Expression and Functional Activity of ABCA4 Disease Variants With the Phenotype of Patients With Stargardt Disease.

Purpose: Stargardt disease (STGD1), the most common early-onset recessive macular degeneration, is caused by mutations in the gene encoding the ATP-binding cassette transporter ABCA4. Although extensive genetic studies have identified more than 1000 mutations that cause STGD1 and related ABCA4-associated diseases, few studies have investigated the extent to which mutations affect the biochemical properties of ABCA4. The purpose of this study was to correlate the expression and functional activities of missense mutations in ABCA4 identified in a cohort of Canadian patients with their clinical phenotype.