Genetic Selection for Enhanced Folding In Vivo Targets the Cys14-Cys38 Disulfide Bond in Bovine Pancreatic Trypsin Inhibitor

The periplasm provides a strongly oxidizing environment; however, periplasmic expression of proteins with disulfide bonds is often inefficient. Here, we used two different tripartite fusion systems to perform in vivo selections for mutants of the model protein bovine pancreatic trypsin inhibitor (BPTI) with the aim of enhancing its expression in Escherichia coli. This trypsin inhibitor contains three disulfides that contribute to its extreme stability and protease resistance. The mutants we isolated for increased expression appear to act by eliminating or destabilizing the Cys14-Cys38 disulfide in BPTI. In doing so, they are expected to reduce or eliminate kinetic traps that exist within the well characterized in vitro folding pathway of BPTI. These results suggest that elimination or destabilization of a disulfide bond whose formation is problematic in vitro can enhance in vivo protein folding. The use of these in vivo selections may prove a valuable way to identify and eliminate disulfides and other rate-limiting steps in the folding of proteins, including those proteins whose in vitro folding pathways are unknown. Antioxid. Redox Signal. 14, 973-984.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/90494/1/ars-2E2010-2E3712.pd

An in vivo platform for identifying inhibitors of protein aggregation

Protein aggregation underlies an array of human diseases, yet only one small molecule therapeutic has been successfully developed to date. Here, we introduce an in vivo system, based on a β-lactamase tripartite fusion construct, capable of identifying aggregation-prone sequences in the periplasm of Escherichia coli and inhibitors that prevent their aberrant self-assembly. We demonstrate the power of the system using a range of proteins, from small unstructured peptides (islet amyloid polypeptide and amyloid β) to larger, folded immunoglobulin domains. Configured in a 48-well format, the split β-lactamase sensor readily differentiates between aggregation-prone and soluble sequences. Performing the assay in the presence of 109 compounds enabled a rank ordering of inhibition and revealed a new inhibitor of IAPP aggregation. This platform can be applied to both amyloidogenic and other aggregation-prone systems, independent of sequence or size, and can identify small molecules or other factors able to ameliorate or inhibit protein aggregation

Management priorities of digital health service start-ups in California

Abstract Digitalisation has revolutionised health service delivery, which has provided global business opportunities for start-ups that specialise in digital innovations. Such start-ups challenge the traditional healthcare service industry by introducing radical and sustainable innovations in the agile product development cycle and the creative acquisition of resources within their networks. Starting up is the most critical period in establishing a new digital health service company. However, little is known about the critical early growth processes of newly established digital healthcare service businesses. The aim of this study is to clarify the experience-based priorities of managers of digital health service businesses in California during the critical start-up stage. Based on this multiple case study, the qualitative and contextual characteristics of growth in California-based digital health service start-ups were clarified, and a framework of management priorities was formed. Network management is a high priority in digital service start-ups that are focused on bringing radical innovations to the complex and hard-to-access market where fundraising is an integral part of success

White Syngas by Co-Electrolysis for Industrial Chemistry

White syngas is produced by co-electrolysis of carbon dioxide and water. As syngas is one of the essential petrochemical foundations of industrial chemistry, the high temperature co-electrolysis can induce a possibility for the defossilization process in conventional petro chemistry. Therefore, it is necessary to discuss the proper technological framework in the scopes of supply of CO2, electrochemical performance of Solid Oxide Electrolysis Cells (SOEC) and products of white syngas. In future Power-to-X scenarios with a 100 % share of renewable energy, it is fundamentally important to calculate process-related carbon dioxide emissions from the total CO2 emissions. Also, we show the superior performance of co-electrolysis to different electrolysis technologies. As a standard of comparison, we introduce the Fossil Carbon Equivalent (FCE) to clarify the impact of white syngas on industrial chemistry by matching energy demand, need of installed electrolysis capacities, consumption of carbon dioxide and substitutable amount of fossil resources