Real-time monitoring of solid-phase peptide synthesis using a variable bed glow reactor

On-resin aggregation and incomplete amide bond formation are major challenges for solid-phase peptide synthesis that are difficult to be monitored in real-time. Incorporation of a pressure-based variable bed flow reactor into an automated solid-phase peptide synthesizer permitted real-time monitoring of resin swelling to determine amino acid coupling efficiency and on-resin aggregation

Towards a systematic understanding of the influence of temperature on glycosylation reactions

Glycosidic bond formation is a continual challenge for practitioners. Aiming to enhance the reproducibility and efficiency of oligosaccharide synthesis, we studied the relationship between glycosyl donor activation and reaction temperature. A novel semi-automated assay revealed diverse responses of members of a panel of thioglycosides to activation at various temperatures. The patterns of protecting groups and the thiol aglycon combine to cause remarkable differences in temperature sensitivity among glycosylating agents. We introduce the concept of donor activation temperature to capture experimental insights, reasoning that glycosylations performed below this reference temperature evade deleterious side reactions. Activation temperatures enable a simplified temperature treatment and facilitate optimization of glycosylating agent (building block) usage. Isothermal glycosylation below the activation temperature halved the equivalents of building block required in comparison to the standard ‘ramp’ regime used in solution- and solid-phase oligosaccharide synthesis to-date

On resin synthesis of sulfated oligosaccharides

Sulfated glycans are involved in many biological processes, making well-defined sulfated oligosaccharides highly sought molecular probes. These compounds are a considerable synthetic challenge, with each oligosaccharide target requiring specific synthetic protocols and extensive purifications steps. Here, we describe a general on resin approach that simplifies the synthesis of sulfated glycans. The oligosaccharide backbone, obtained by Automated Glycan Assembly (AGA), is subjected to regioselective sulfation and hydrolysis of protecting groups. The protocol is compatible with several monosaccharides and allows for multi-sulfation of linear and branched glycans. Seven diverse, biologically relevant sulfated glycans were prepared in good to excellent overall yield

Guidance on marine protected area protection level assignments when faced with unknown regulatory information

Strong human use regulations are an important precondition for marine protected area (MPA) effectiveness. Distinguishing MPAs based on their protection levels has shown advantages, but the availability of regulatory information about allowed activities is a major roadblock towards completing assessments at scale. Here, using a California case study, we explore assigning MPA protection levels following the regulation-based classification system (RBCS) under different scenarios of incomplete regulatory information. In the first group of scenarios (A), only readily available information was used, i.e., information contained in direct MPA implementing regulations and management plans. In the second group (B), information was limited to the activities in ProtectedSeas' Navigator that matched those in the RBCS. From group A, 99% and 100% correct classification of fully and highly protected areas, respectively, were obtained when treating unknown aquaculture, bottom exploitation, and bottom extraction as 'prohibited' and boating, anchoring, and fishing activities as 'allowed'. High classifi-cation accuracy was also obtained for moderately, poorly, and unprotected areas. From group B, 92% and 94% correct classification of fully and highly protected areas were obtained when using the same assumptions for non -fishing activities but using Navigator's Level of Fishing Protection (LFP) score to guide assumptions about un-known fishing activities. Correct classification rates were poorer with different assumptions. Regulation-based MPA evaluation systems can reliably identify fully and highly protected areas in the face of unknown infor-mation, when assumptions about unknown information are guided by contextual indicators such as generally regulated human activities and/or overall level of fishing restriction.info:eu-repo/semantics/publishedVersio

Microwave-assisted automated glycan assembly

Automated synthesis of DNA, RNA, and peptides provides quickly and reliably important tools for biomedical research. Automated glycan assembly (AGA) is significantly more challenging as highly branched carbohydrates require strict regio- and stereocontrol during synthesis. A new AGA synthesizer enables rapid temperature adjustment from -40 °C to +100 °C to control glycosylations at low temperature and accelerates capping, protecting group removal, and glycan modifications by using elevated temperatures. Thereby, the temporary protecting group portfolio is extended from two to four orthogonal groups that give rise to oligosaccharides with up to four branches. In addition, sulfated glycans and unprotected glycans can be prepared. The new design reduces the typical coupling cycles from 100 min to 60 min while expanding the range of accessible glycans. The instrument drastically shorten and generalizes the synthesis of carbohydrates for use in biomedical and material science

VaporSPOT : parallel synthesis of oligosaccharides on membranes

Automated chemical synthesis has revolutionized synthetic access to biopolymers in terms of simplicity and speed. While automated oligosaccharide synthesis has become faster and more versatile, the parallel synthesis of oligosaccharides is not yet possible. Here, a chemical vapor glycosylation strategy (VaporSPOT) is described that enables the simultaneous synthesis of oligosaccharides on a cellulose membrane solid support. Different linkers allow for flexible and straightforward cleavage, purification, and characterization of the target oligosaccharides. This method is the basis for the development of parallel automated glycan synthesis platforms

Molecular and biogeochemical evidence for methane cycling beneath the western margin of the Greenland Ice Sheet

Microbial processes that mineralize organic carbon and enhance solute production at the bed of polar ice sheets could be of a magnitude sufficient to affect global elemental cycles. To investigate the biogeochemistry of a polar subglacial microbial ecosystem, we analyzed water discharged during the summer of 2012 and 2013 from Russell Glacier, a land-terminating outlet glacier at the western margin of the Greenland Ice Sheet. The molecular data implied that the most abundant and active component of the subglacial microbial community at these marginal locations were bacteria within the order Methylococcales (59–100% of reverse transcribed (RT)-rRNA sequences). mRNA transcripts of the particulate methane monooxygenase (pmoA) from these taxa were also detected, confirming that methanotrophic bacteria were functional members of this subglacial ecosystem. Dissolved methane ranged between 2.7 and 83 μM in the subglacial waters analyzed, and the concentration was inversely correlated with dissolved oxygen while positively correlated with electrical conductivity. Subglacial microbial methane production was supported by δ(13)C-CH(4) values between −64‰ and −62‰ together with the recovery of RT-rRNA sequences that classified within the Methanosarcinales and Methanomicrobiales. Under aerobic conditions, >98% of the methane in the subglacial water was consumed over ∼30 days incubation at ∼4 °C and rates of methane oxidation were estimated at 0.32 μM per day. Our results support the occurrence of active methane cycling beneath this region of the Greenland Ice Sheet, where microbial communities poised in oxygenated subglacial drainage channels could serve as significant methane sinks