Towards understanding the interaction between oligosaccharides and water molecules

Complex carbohydrates are implicated in many important biological processes, and have a strong interaction with water. This close interplay with molecular water through multiple hydroxyls may be an integral part of their emergent structure and dynamics, as selected during evolution. Using molecular dynamics simulations with explicit water the interactions at the linkages within a variety of oligosaccharides are investigated and contrasted, in order to establish correlations between linkage orientation, sugar epimerization, and water interaction. In particular, interactions at alpha linkages, and between mannose and glucose residues, that are common in oligosaccharides are considered. Sugars joined by alpha linkages at the 2-, 3-, and 6-position were found to interact via a combination of weak hydrogen-bonds and water-bridges, which is dependent on the epimerization state of the sugars. Due to their three-dimensional structure, they are also likely to interact with noncontiguous sugar residues in an oligosaccharide, which can lead to ordered structures through the exclusion of water. On the other hand, beta linkages (to 3- and 4-position) maintain strong hydrogen-bonds, have a limited ability to be involved in water-bridges, and predominantly interact with the directly attached sugars. Therefore, sequences of alpha-linked sugars form compact, branched structures that have conformational flexibility, and beta linkages form extended, relatively rigid structures, suitable for structural molecules, and at the termini of protein bound oligosaccharides. These results provide further tentative ties between chemical structure, water interactions, and the emergent form and function of specific sugars and linkages in oligosaccharides.     

The immunohistochemical location of cathepsin L in rabbit skeletal muscle. Evidence for a fibre type dependent distribution

The immunohistochemical location of cathepsin L in rabbit soleus, plantaris and psoas muscles was investigated using the peroxidase-anti-peroxidase (PAP) technique. The amount of enzyme detected varied according to the fibre type, which were identified by histochemical staining of serial sections for succinate dehydrogenase and alkali-stable myosin ATPase. In the three muscles studied labelling was strongest in the highly oxidative fibres and weaker in the other fibre types with least staining in the fast white fibres. Immunoreactive cathepsin L appeared to be most concentrated at the periphery of muscle fibres, especially near to the nuclei, although some staining was seen throughout the fibres.     

Horizontal acquisition and a broad biodistribution typify simian foamy virus infection in a cohort of Macaca fascicularis

Bluetongue virus (BTV) is an arbovirus that is responsible for ‘bluetongue’, an economically important disease of livestock. Although BTV is well characterised at the protein level, less is known regarding its interaction with host cells. During studies of virus inclusion body formation we observed what appeared to be a large proportion of cells in mitosis. Although the modulation of the cell cycle is well established for many viruses, this was a novel observation for BTV. We therefore undertook a study to reveal in more depth the impact of BTV upon cell division. We used a confocal microscopy approach to investigate the localisation of BTV proteins in a cellular context with their respective position relative to cellular proteins. In addition, to quantitatively assess the frequency of aberrant mitosis induction by the viral non-structural protein (NS) 2 we utilised live cell imaging to monitor HeLa-mCherry tubulin cells transfected with a plasmid expressing NS2. Our data showed that these ‘aberrant mitoses’ can be induced in multiple cell types and by different strains of BTV. Further study confirmed multiplication of the centrosomes, each resulting in a separate mitotic spindle during mitosis. Interestingly, the BTV NS1 protein was strongly localised to the centrosomal regions. In a separate, yet related observation, the BTV NS2 protein was co-localised with the condensed chromosomes to a region suggestive of the kinetochore. Live cell imaging revealed that expression of an EGFP-NS2 fusion protein in HeLa-mCherry tubulin cells also results in mitotic defects. We hypothesise that NS2 is a microtubule cargo protein that may inadvertently disrupt the interaction of microtubule tips with the kinetochores during mitosis. Furthermore, the BTV NS1 protein was distinctly localised to a region encompassing the centrosome and may therefore be, at least in part, responsible for the disruption of the centrosome as observed in BTV infected mammalian cells.     

Enzymatic and chemical methods for the generation of pure hyaluronan oligosaccharides with both odd and even numbers of monosaccharide units

Hyaluronan oligosaccharides display physiological activities not associated with the polymer and are widely used to characterize hyaluronan-binding proteins. They can also be used as biocompatible starting blocks for chemical derivatization. Here we present methods for generating milligram quantities of unusual odd- and even-numbered oligosaccharides, greatly increasing the diversity of reagents for use in such studies. These methods are based upon protocols from the 1960s, at which time it was very difficult to assess the stereochemical purity of the products. To address this, products were analyzed with modern high-field nuclear magnetic resonance spectroscopy. Alkaline beta-elimination conditions previously used to remove reducing-terminal N-acetylglucosamine residues in fact introduce a significant ( approximately 30%) level of stereoisomerism in the products by alkali-catalyzed keto-enol tautomerizations. Milder alkaline conditions were used to overcome this problem, reducing the contamination to <5%. The elimination by-products from this reaction were isolated and characterized, allowing the mechanism of alkaline degradation of hyaluronan to be investigated for the first time. beta-Glucuronidase was used to remove nonreducing-terminal glucuronic acid residues from oligosaccharides. Odd-numbered oligosaccharides with terminal glucuronic acid residues isolated from hyaluronidase digests are shown to originate from acid-catalyzed acetal hydrolysis during boiling denaturation and also have significant levels of stereochemical impurities.     

Genetic profiling of newborns: ethical and social issues

Identifying genetic factors that could reliably predict health risks for individuals has the potential to bring great health benefits, both for the individuals concerned and for health-care providers. Genetic profiling at birth would allow a person's genome to be analysed at an early stage, and the data electronically stored for future use. However, although this might seem like an attractive proposition, it carries with it serious ethical and social concerns that would need to be addressed if the genetic profiling of newborns were ever to be considered on a population-wide basis.     

Discovery Proteomics Analysis Determines That Driver Oncogenes Suppress Antiviral Defense Pathways Through Reduction in Interferon-β Autocrine Stimulation

Since the discovery of oncogenes, there has been tremendous interest to understand their mechanistic basis and to develop broadly actionable therapeutics. Some of the most frequently activated oncogenes driving diverse cancers are c-MYC, EGFR, HER2, AKT, KRAS, BRAF, and MEK. Using a reductionist approach, we explored how cellular proteomes are remodeled in isogenic cell lines engineered with or without these driver oncogenes. The most striking discovery for all oncogenic models was the systematic downregulation of scores of antiviral proteins regulated by type 1 interferon. These findings extended to cancer cell lines and patient-derived xenograft models of highly refractory pancreatic cancer and osteosarcoma driven by KRAS and MYC oncogenes. The oncogenes reduced basal expression of and autocrine stimulation by type 1 interferon causing remarkable convergence on common phenotypic and functional profiles. In particular, there was dramatically lower expression of dsRNA sensors including DDX58 (RIG-I) and OAS proteins, which resulted in attenuated functional responses when the oncogenic cells were treated with the dsRNA mimetic, polyI:C, and increased susceptibility to infection with an RNA virus shown using SARS-CoV-2. Our reductionist approach provides molecular and functional insights connected to immune evasion hallmarks in cancers and suggests therapeutic opportunities.     

Novel inhibitors of HIV protease: design, synthesis and biological evaluation of picomolar inhibitors containing cyclic P1/P2 scaffolds

A novel series of HIV protease inhibitors containing cyclic P1/P2 scaffolds has been synthesized and evaluated for biological activity. The trans 3,5-dibenzyl-2-oxo pyrrolidinone ring system resulted in a 50 pM enzyme inhibitor against HIV protease in vitro when combined with an indanolamine derived P'-backbone. This compound also shows comparable activity to currently marketed drugs in the MT-4 cell-based antiviral assay.     

Modeled distributions of Aedes japonicus japonicus and Aedes togoi (Diptera: Culicidae) in the United States,Canada, and northern Latin America

The Asian bush mosquito, Aedes japonicus japonicus, and the coastal rock pool mosquito, Aedes togoi, are potential disease vectors present in both East Asia and North America. While their ranges are fairly well‐documented in Asia, this is not the case for North America. We used maximum entropy modeling to estimate the potential distributions of Ae. togoi and Ae. j. japonicus in the United States, Canada, and northern Latin America under contemporary and future climatic conditions. Our results suggest suitable habitat that is not known to be occupied for Ae. j. japonicus in Atlantic and western Canada, Alaska, the western, midwestern, southern, and northeastern United States, and Latin America, and for Ae. togoi along the Pacific coast of North America and the Hawaiian Islands. Such areas are at risk of future invasion or may already contain undetected populations of these species. Our findings further predict that the limits of suitable habitat for each species will expand northward under future climatic conditions.