Neutralization potency of monoclonal antibodies recognizing dominant and subdominant epitopes on SARS-CoV-2 Spike is impacted by the B.1.1.7 variant

Interaction of the SARS-CoV-2 Spike receptor binding domain (RBD) with the receptor ACE2 on host cells is essential for viral entry. RBD is the dominant target for neutralizing antibodies, and several neutralizing epitopes on RBD have been molecularly characterized. Analysis of circulating SARS-CoV-2 variants has revealed mutations arising in the RBD, N-terminal domain (NTD) and S2 subunits of Spike. To understand how these mutations affect Spike antigenicity, we isolated and characterized >100 monoclonal antibodies targeting epitopes on RBD, NTD, and S2 from SARS-CoV-2-infected individuals. Approximately 45% showed neutralizing activity, of which ∼20% were NTD specific. NTD-specific antibodies formed two distinct groups: the first was highly potent against infectious virus, whereas the second was less potent and displayed glycan-dependant neutralization activity. Mutations present in B.1.1.7 Spike frequently conferred neutralization resistance to NTD-specific antibodies. This work demonstrates that neutralizing antibodies targeting subdominant epitopes should be considered when investigating antigenic drift in emerging variants

Applications of isothermal titration calorimetry - the research and technical developments from 2011 to 2015

Isothermal titration calorimetry is a widely used biophysical technique for studying the formation or dissociation of molecular complexes. Over the last 5years, much work has been published on the interpretation of isothermal titration calorimetry (ITC) data for single binding and multiple binding sites. As over 80% of ITC papers are on macromolecules of biological origin, this interpretation is challenging. Some researchers have attempted to link the thermodynamics constants to events at the molecular level. This review highlights work carried out using binding sites characterized using x-ray crystallography techniques that allow speculation about individual bond formation and the displacement of individual water molecules during ligand binding and link these events to the thermodynamic constants for binding. The review also considers research conducted with synthetic binding partners where specific binding events like anion-π and π-π interactions were studied. The revival of assays that enable both thermodynamic and kinetic information to be collected from ITC data is highlighted. Lastly, published criticism of ITC research from a physical chemistry perspective is appraised and practical advice provided for researchers unfamiliar with thermodynamics and its interpretation

Experiences of health care providers with integrated HIV and reproductive health services in Kenya: a qualitative study.

BACKGROUND: There is broad consensus on the value of integration of HIV services and reproductive health services in regions of the world with generalised HIV/AIDS epidemics and high reproductive morbidity. Integration is thought to increase access to and uptake of health services; and improves their efficiency and cost-effectiveness through better use of available resources. However, there is still very limited empirical literature on health service providers and how they experience and operationalize integration. This qualitative study was conducted among frontline health workers to explore provider experiences with integration in order to ascertain their significance to the performance of integrated health facilities. METHODS: Semi-structured in-depth interviews were conducted with 32 frontline clinical officers, registered nurses, and enrolled nurses in Kitui district (Eastern province) and Thika and Nyeri districts (Central province) in Kenya. The study was conducted in health facilities providing integrated HIV and reproductive health services (post-natal care and family planning). All interviews were conducted in English, transcribed and analysed using Nvivo 8 qualitative data analysis software. RESULTS: Providers reported delivering services in provider-level and unit-level integration, as well as a combination of both. Provider experiences of actual integration were mixed. At personal level, providers valued skills enhancement, more variety and challenge in their work, better job satisfaction through increased client-satisfaction. However, they also felt that their salaries were poor, they faced increased occupational stress from: increased workload, treating very sick/poor clients, and less quality time with clients. At operational level, providers reported increased service uptake, increased willingness among clients to take an HIV test, and reduced loss of clients. But the majority also reported infrastructural and logistic deficiencies (insufficient physical room space, equipment, drugs and other medical supplies), as well as increased workload, waiting times, contact session times and low staffing levels. CONCLUSIONS: The success of integration primarily depends on the performance of service providers which, in turn, depends on a whole range of facilitative organisational factors. The central Ministry of Health should create a coherent policy environment, spearhead strategic planning and ensure availability of resources for implementation at lower levels of the health system. Health facility staffing norms, technical support, cost-sharing policies, clinical reporting procedures, salary and incentive schemes, clinical supply chains, and resourcing of health facility physical space upgrades, all need attention. Yet, despite these system challenges, this study has shown that integration can have a positive motivating effect on staff and can lead to better sharing of workload - these are important opportunities that deserve to be built on

The ABC transporter BmrA from Bacillus subtilis is a functional dimer when in a detergent-solubilized state

BmrA from Bacillus subtilis is a half-size ABC (ATP-binding cassette) transporter involved in multidrug resistance. Although its supramolecular organization has been investigated after reconstitution in a lipid bilayer environment, and shows a dimeric and possibly a tetrameric form, the precise quaternary structure in a detergent-solubilized state has never been addressed. In the present study, BmrA was purified from Escherichia coli membranes using an optimized purification protocol and different detergents. Furthermore, the ATPase activity of BmrA and the quantity of bound lipids and detergent were determined, and the oligomeric state was analysed using SEC (size-exclusion chromatography) and analytical ultracentrifugation. The activity and the quaternary structure of BmrA appeared to be strongly influenced by the type and concentration of the detergent used. SEC data showed that BmrA could be purified in a functional form in 0.05 and 0.01% DDM (n-dodecyl-β-D-maltoside) and was homogeneous and monodisperse with an R(s) (Stokes radius) of 5.6 nm that is compatible with a dimer structure. Sedimentation-velocity and equilibrium experiments unequivocally supported that BmrA purified in DDM is a dimer and excluded the presence of other oligomeric states. These observations, which are discussed in relation to results obtained in proteoliposomes, also constitute an important first step towards crystallographic studies of BmrA structure