Recombinant enzyme expression in Escherichia coli is one of the most popular methods to produce bulk concentrations of protein product. However, this method is often limited by the inadvertent formation of inclusion bodies. Our analysis systematically reviews literature from 2010 to 2021 and details the methods and strategies researchers have utilized for expression of difficult to express (DtE), industrially relevant recombinant enzymes in E. coli expression strains. Our review identifies an absence of a coherent strategy with disparate practices being used to promote solubility. We discuss the potential to approach recombinant expression systematically, with the aid of modern bioinformatics, modelling, and ‘omics’ based systems-level analysis techniques to provide a structured, holistic approach. Our analysis also identifies potential gaps in the methods used to report metadata in publications and the impact on the reproducibility and growth of the research in this field.

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release. By rational design of its core structure to alter (photo)physical properties, we obtained derivatives capable of outperforming riboflavin’s visible light-induced PDI against E. coli and a SARS-CoV-2 surrogate, revealing functional group dependency for each pathogen. Bacterial PDI was influenced mainly by guanidino substitution, whereas viral PDI increased through bromination of the flavin. These observations were related to enhanced uptake and ROS-specific nucleic acid cleavage mechanisms. Trends in the derivatives’ toxicity towards human fibroblast cells were also investigated to assess viable therapeutic derivatives and help guide further design of PDI agents to combat pathogenic organisms.

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release. By rational design of its core structure to alter (photo)physical properties, we obtained derivatives capable of outperforming riboflavin’s visible light-iinduced PDI against E. coli and a SARS-CoV-2 surrogate, revealing functional group dependency for each pathogen. Bacterial PDI was influenced mainly by guanidino substitution, whereas viral PDI increased through bromination of the flavin. These observations were related to enhanced uptake and ROS-specific nucleic acid cleavage mechanisms. Trends in the derivatives’ toxicity towards human fibroblast cells were also investigated to assess viable therapeutic derivatives and help guide further design of PDI agents to combat pathogenic organisms.

The development of effective pathogen reduction strategies is required due to the rise in antibiotic-resistant bacteria and zoonotic viral pandemics. Photodynamic inactivation (PDI) of bacteria and viruses is a potent reduction strategy that bypasses typical resistance mechanisms. Naturally occurring riboflavin has been widely used in PDI applications due to efficient light-induced reactive oxygen species (ROS) release. By rational design of its core structure to alter (photo)physical properties, we obtained derivatives capable of outperforming riboflavin’s visible light-iinduced PDI against E. coli and a SARS-CoV-2 surrogate, revealing functional group dependency for each pathogen. Bacterial PDI was influenced mainly by guanidino substitution, whereas viral PDI increased through bromination of the flavin. These observations were related to enhanced uptake and ROS-specific nucleic acid cleavage mechanisms. Trends in the derivatives’ toxicity towards human fibroblast cells were also investigated to assess viable therapeutic derivatives and help guide further design of PDI agents to combat pathogenic organisms.