Biology subjects

Bond, P.

Publications and source records attributed to Bond, P..

3 recordsLinked to original sources

bioRxiv2019

Non-antibiotic pharmaceuticals can enhance the spread of antibiotic resistance via conjugation

Antibiotic resistance is a global threat for public health. It is widely acknowledged that antibiotics at sub-inhibitory concentrations are important in disseminating antibiotic resistance via horizontal gene transfer. While there is high use of non-antibiotic human-targeted pharmaceuticals in our societies, the potential contribution of these on the spread of antibiotic resistance has been overlooked so far. Here, we report that commonly consumed non-antibiotic pharmaceuticals, including nonsteroidal anti-inflammatories (ibuprofen, naproxen, diclofenac), a lipid-lowering drug (gemfibrozil), and a {beta}-blocker (propanolol), at clinically and environmentally relevant concentrations, significantly accelerated the conjugation of plasmid-borne antibiotic resistance genes. We looked at the response to these drugs by the bacteria involved in the gene transfer through various analyses that included monitoring reactive oxygen species (ROS) and cell membrane permeability by flow cytometry, cell arrangement, and whole-genome RNA and protein sequencing. We found the enhanced conjugation correlated well with increased production of ROS and cell membrane permeability. We also detected closer cell-to-cell contact and upregulated conjugal genes. Additionally, these non-antibiotic pharmaceuticals caused the bacteria to have responses similar to those detected when exposed to antibiotics, such as inducing the SOS response, and enhancing efflux pumps. The findings advance our understanding of the bacterial transfer of antibiotic resistance genes, and importantly emphasize concerns of non-antibiotic human-targeted pharmaceuticals for enhancing the spread of antibiotic resistance.

microbiology↗

bioRxiv2019

The structural basis for distinct binding avidity of Pertuzumab and Trastuzumab IgM towards HER2

Harnessing the therapeutic potential of immunoglobulin M (IgM) is of considerable interest in immunotherapy due to its complement-activating and cell-agglutinating abilities. Pertuzumab and Trastuzumab are monoclonal antibody drugs used in therapy for patients with human epidermal growth factor receptor 2 (HER2)-positive breast cancer but exhibit significantly different binding affinities as IgM when compared to the original IgG1 form. While the affinity of Pertuzumab IgM to the HER2 extracellular domain is about one order of magnitude higher than IgG1 in experiments, it was recently reported that Trastuzumab IgM and IgG have similar equilibrium dissociation constants to one another. We now perform an integrative multiscale simulation study in order to understand the structural basis for the differences in behavior between the two antibodies, based on complete antibody assemblies. We show that Pertuzumab IgM is able to utilize all of its V-regions to engage HER2 in a more stable mode than Trastuzumab IgM due to steric clashes between the large globular HER2 domains when bound to Trastuzumab. This is subsequently validated by confirming that Pertuzumab IgM inhibits proliferation in HER2 over-expressing live cells more effectively than its IgG1 counterpart. Given the widespread clinical use of Trastuzumab and Pertuzumab, elucidating the molecular details of antibody-antigen interaction may help guide the choice of epitopes for future design and selection of improved therapeutic antibody isotypes.

immunology↗

bioRxiv2019

Genome-wide Associations of Flavivirus Capsid Proteins

Dengue virus (DENV) and Zika virus (ZIKV) are both positive sense single-stranded RNA viruses. They are packaged within the virion with a capsid (C) protein to form the nucleocapsid. Based on cryo-electron microscopy imaging, the nucleocapsid has been described as lacking symmetry, whilst there is distinguishable separation of the C proteins from the viral RNA (vRNA) genome. Here, to elucidate the architecture of the nucleocapsid of DENV serotype 2 and ZIKV, we used a nuclease digestion assay and next-generation sequencing to map the respective vRNA genome wide association with the C protein in vitro. The C protein exhibited non-uniform binding along the vRNA, and as C protein concentration increased, the normalized read counts also increased. A saturation point of 1:100 (vRNA:C protein monomers) was found, and binding regions showed variable saturation patterns. We also observed that C protein had a preference for G-rich sequences for both viruses. Taken together, we demonstrate that the DENV 2 and ZIKV C proteins bind vRNA in a non-uniform manner with distinct patterns of association.\n\nSingificance StatementOur study demonstrates that flavivirus capsid proteins associate with the viral genome at specific sites rather than in a uniform manner as commonly expected. We estimate the number of capsid proteins binding to a single genomic RNA. We proceed to locate the capsid binding sites along the viral genomes of Dengue and Zika viruses. We characterize the binding sites in terms of affinity and analyze the nucleotide composition and sequence motifs at binding sites. We cross-reference binding sites against SHAPE reactivity data corresponding to local RNA secondary structure, which allows us to identify structural motifs of capsid binding sites. As capsid proteins are essential for viral packaging, these interactions may form attractive targets for therapeutic intervention.

genomics↗