Coupling and Activation of the β1 Adrenergic Receptor - The Role of the Third Intracellular Loop.

G protein-coupled receptors (GPCRs) belong to the most diverse group of membrane receptors with a conserved structure of seven transmembrane (TM) α-helices connected by intracellular and extracellular loops. Intracellular loop 3 (ICL3) connects TM5 and TM6, the two helices shown to play significant roles in receptor activation. Herein, we investigate the activation and signaling of the β adrenergic receptor (βAR) using mass spectrometry (MS) with a particular focus on the ICL3 loop.

Coarse-Grained Simulations of Adeno-Associated Virus and Its Receptor Reveal Influences on Membrane Lipid Organization and Curvature.

Adeno-associated virus (AAV) is a well-known gene delivery tool with a wide range of applications, including as a vector for gene therapies. However, the molecular mechanism of its cell entry remains unknown. Here, we performed coarse-grained molecular dynamics simulations of the AAV serotype 2 (AAV2) capsid and the universal AAV receptor (AAVR) in a model plasma membrane environment.

Lipid regulation of the glucagon receptor family.

The glucagon receptor family are typical class B1 G protein-coupled receptors (GPCRs) with important roles in metabolism, including the control of pancreas, brain, and liver function. As proteins with seven transmembrane domains, GPCRs are intimately in contact with lipid bilayers and therefore can be putatively regulated by interactions with their lipidic components, including cholesterol, sphingolipids, and other lipid species. Additionally, these receptors, as well as the agonists they bind to, can undergo lipid modifications, which can influence their binding capacity and/or elicit modified or biased signalling profiles.

The protease associated (PA) domain in ScpA from Streptococcus pyogenes plays a role in substrate recruitment.

Annually, over 18 million disease cases and half a million deaths worldwide are estimated to be caused by Group A Streptococcus. ScpA (or C5a peptidase) is a well characterised member of the cell enveleope protease family, which possess a S8 subtilisin-like catalytic domain and a shared multi-domain architecture. ScpA cleaves complement factors C5a and C3a, impairing the function of these critical anaphylatoxins and disrupts complement-mediated innate immunity.

Structural and regulatory insights into the glideosome-associated connector from .

The phylum of Apicomplexa groups intracellular parasites that employ substrate-dependent gliding motility to invade host cells, egress from the infected cells, and cross biological barriers. The glideosome-associated connector (GAC) is a conserved protein essential to this process. GAC facilitates the association of actin filaments with surface transmembrane adhesins and the efficient transmission of the force generated by myosin translocation of actin to the cell surface substrate.

Measurement of Adenovirus-Based Vector Heterogeneity.

Adenovirus vectors have become an important class of vaccines with the recent approval of Ebola and COVID-19 products. In-process quality attribute data collected during Adenovirus vector manufacturing has focused on particle concentration and infectivity ratios (based on viral genome: cell-based infectivity), and data suggest only a fraction of viral particles present in the final vaccine product are efficacious. To better understand this product heterogeneity, lab-scale preparations of two Adenovirus viral vectors, (Chimpanzee adenovirus (ChAdOx1) and Human adenovirus Type 5 (Ad5), were studied using transmission electron microscopy (TEM).

An intermolecular hydrogen bonded network in the PRELID-TRIAP protein family plays a role in lipid sensing.

The PRELID-TRIAP1 family of proteins is responsible for lipid transfer in mitochondria. Multiple structures have been resolved of apo and lipid substrate bound forms, allowing us to begin to piece together the molecular level details of the full lipid transfer cycle. Here, we used molecular dynamics simulations to demonstrate that the lipid binding is mediated by an extended, water-mediated hydrogen bonding network.

Corrigendum to "Atomistic level characterisation of ssDNA translocation through the proteins CsgG and CsgF for nanopore sequencing" [Comput. Struct. Biotechnol. J. 19 (2021) 6417-6430].

[This corrects the article DOI: 10.1016/j.csbj.

Intraspecific interactions in a high-density leopard population.

Although less studied than interspecific interactions, interactions among members of the same species can influence space use and temporal activity. Using techniques commonly applied to the analysis of interspecific interactions-multispecies occupancy modeling and the analysis of temporal activity patterns-we studied intraspecific interactions within a high-density population of Persian leopards () in Tandoureh National Park, northeastern Iran. Using camera-trap data, we investigated spatiotemporal interactions between male leopards, lone female leopards, and families (cubs/females with cubs).

Atomistic level characterisation of ssDNA translocation through the proteins CsgG and CsgF for nanopore sequencing.

Two proteins of the membrane protein complex, CsgG and CsgF, are studied as proteinaceous nanopores for DNA sequencing. It is highly desirable to control the DNA as it moves through the pores, this requires characterisation of DNA translocation and subsequent optimization of the pores. In order to inform protein engineering to improve the pores, we have conducted a series of molecular dynamics simulations to characterise the mechanical strength and conformational dynamics of CsgG and the CsgG-CsgF complex and how these impact ssDNA, water and ion movement.

Structure of the cytoplasmic domain of SctV (SsaV) from the Salmonella SPI-2 injectisome and implications for a pH sensing mechanism.

Bacterial type III secretion systems assemble the axial structures of both injectisomes and flagella. Injectisome type III secretion systems subsequently secrete effector proteins through their hollow needle into a host, requiring co-ordination. In the Salmonella enterica serovar Typhimurium SPI-2 injectisome, this switch is triggered by sensing the neutral pH of the host cytoplasm.

Caustic ingestions over 10 years in Victoria, Australia: high rates in migrants and women.

Background: Caustic ingestion is relatively common in developing countries and can result in life-threatening sequelae. There is limited understanding of the epidemiology and incidence in Australia.

Aims: This statewide 10-year audit aims to document the rate of caustic injury in a defined Australian pouplation.

Does Previous Cardiac Surgery Predict Impaired Quality of Life in Adults With Congenital Heart Disease?

Background: Improved survival of children with congenital heart disease (CHD) into adult life has led to further study of their quality of life (QoL) and its determinants. The QoL including the symptoms of anxiety and depression of adults with CHD was analyzed to determine the relationship, if any, between prior cardiac surgery and QoL.

Methods: Adults with CHD who were recruited from a single community-based cardiology practice completed self-reported questionnaires on their QoL, which included symptoms of anxiety and depression.

Structure, dynamics and immunogenicity of a catalytically inactive CC chemokine-degrading protease SpyCEP from .

Over 18 million disease cases and half a million deaths worldwide are estimated to be caused annually by Group A Streptococcus. A vaccine to prevent GAS disease is urgently needed. SpyCEP (Streptococcus Cell-Envelope Proteinase) is a surface-exposed serine protease that inactivates chemokines, impairing neutrophil recruitment and bacterial clearance, and has shown promising immunogenicity in preclinical models.

The use of sonicated lipid vesicles for mass spectrometry of membrane protein complexes.

Recent applications of mass spectrometry (MS) to study membrane protein complexes are yielding valuable insights into the binding of lipids and their structural and functional roles. To date, most native MS experiments with membrane proteins are based on detergent solubilization. Many insights into the structure and function of membrane proteins have been obtained using detergents; however, these can promote local lipid rearrangement and can cause fluctuations in the oligomeric state of protein complexes.

Simulation of subcellular structures.

Advances in molecular dynamics simulations have led to large increases across spatial and complexity scales, providing valuable molecular level insight into processes occurring on the subcellular level. An increasing repertoire of methods to assemble and analyse complex membrane simulations, alongside advances in structural biology methods for membrane proteins, have contributed to our increased understanding of the roles of specific lipid interactions for multiple membrane protein systems. Large scale simulations of crowded protein solutions have provided a model describing the biophysical basis for experimentally observed diffusion properties.

Structural determinants of lipid specificity within Ups/PRELI lipid transfer proteins.

Conserved lipid transfer proteins of the Ups/PRELI family regulate lipid accumulation in mitochondria by shuttling phospholipids in a lipid-specific manner across the intermembrane space. Here, we combine structural analysis, unbiased genetic approaches in yeast and molecular dynamics simulations to unravel determinants of lipid specificity within the conserved Ups/PRELI family. We present structures of human PRELID1-TRIAP1 and PRELID3b-TRIAP1 complexes, which exert lipid transfer activity for phosphatidic acid and phosphatidylserine, respectively.

Protein assemblies ejected directly from native membranes yield complexes for mass spectrometry.

Membrane proteins reside in lipid bilayers and are typically extracted from this environment for study, which often compromises their integrity. In this work, we ejected intact assemblies from membranes, without chemical disruption, and used mass spectrometry to define their composition. From outer membranes, we identified a chaperone-porin association and lipid interactions in the β-barrel assembly machinery.

Structural Basis of Phosphatidic Acid Sensing by APH in Apicomplexan Parasites.

Plasmodium falciparum and Toxoplasma gondii are obligate intracellular parasites that belong to the phylum of Apicomplexa and cause major human diseases. Their access to an intracellular lifestyle is reliant on the coordinated release of proteins from the specialized apical organelles called micronemes and rhoptries. A specific phosphatidic acid effector, the acylated pleckstrin homology domain-containing protein (APH) plays a central role in microneme exocytosis and thus is essential for motility, cell entry, and egress.

The FapF Amyloid Secretion Transporter Possesses an Atypical Asymmetric Coiled Coil.

Gram-negative bacteria possess specialized biogenesis machineries that facilitate the export of amyloid subunits, the fibers of which are key components of their biofilm matrix. The secretion of bacterial functional amyloid requires a specialized outer-membrane protein channel through which unfolded amyloid substrates are translocated. We previously reported the crystal structure of the membrane-spanning domain of the amyloid subunit transporter FapF from Pseudomonas.

Ecology and Biogenesis of Functional Amyloids in Pseudomonas.

Functional amyloids can be found in the extracellular matrix produced by many bacteria during biofilm growth. They mediate the initial attachment of bacteria to surfaces and provide stability and functionality to mature biofilms. Efficient amyloid biogenesis requires a highly coordinated system of amyloid subunits, molecular chaperones and transport systems.

A new class of hybrid secretion system is employed in Pseudomonas amyloid biogenesis.

Gram-negative bacteria possess specialised biogenesis machineries that facilitate the export of amyloid subunits for construction of a biofilm matrix. The secretion of bacterial functional amyloid requires a bespoke outer-membrane protein channel through which unfolded amyloid substrates are translocated. Here, we combine X-ray crystallography, native mass spectrometry, single-channel electrical recording, molecular simulations and circular dichroism measurements to provide high-resolution structural insight into the functional amyloid transporter from Pseudomonas, FapF.

Purification, crystallization and characterization of the Pseudomonas outer membrane protein FapF, a functional amyloid transporter.

Bacteria often produce extracellular amyloid fibres via a multi-component secretion system. Aggregation-prone, unstructured subunits cross the periplasm and are secreted through the outer membrane, after which they self-assemble. Here, significant progress is presented towards solving the high-resolution crystal structure of the novel amyloid transporter FapF from Pseudomonas, which facilitates the secretion of the amyloid-forming polypeptide FapC across the bacterial outer membrane.

Structural insights into functional amyloid inhibition in Gram -ve bacteria.

Amyloids are proteinaceous aggregates known for their role in debilitating degenerative diseases involving protein dysfunction. Many forms of functional amyloid are also produced in nature and often these systems require careful control of their assembly to avoid the potentially toxic effects. The best-characterised functional amyloid system is the bacterial curli system.