Propane and butane are the main constituents of liquefied petroleum gas and are used extensively for transport and domestic use. They are clean burning fuels, suitable for the development of low carbon footprint fuel and energy policies. Here, we present blueprints for the production of bio-alkane gas (propane and butane) through the conversion of waste volatile fatty acids by bacterial culture. We show that bio-propane and bio-butane can be produced photo-catalytically by bioengineered strains of E. coli and Halomonas (in non-sterile seawater) using fatty acids derived from biomass or industrial waste, and by Synechocystis (using carbon dioxide as feedstock). Scaled production using available infrastructure is calculated to be economically feasible using Halomonas. These fuel generation routes could be deployed rapidly, in both advanced and developing countries, and contribute to energy security to meet global carbon management targets and clean air directives.

Electron transferring flavoproteins (ETFs) are soluble heterodimeric FAD‐containing proteins that function primarily as soluble electron carriers between various flavoprotein dehydrogenases. ETF is positioned at a key metabolic branch point, responsible for transferring electrons from up to 10 primary dehydrogenases to the membrane‐bound respiratory chain. Clinical mutations of ETF result in the often fatal disease glutaric aciduria type II. Structural and biophysical studies of ETF in complex with partner proteins have shown that ETF partitions the functions of partner binding and electron transfer between (a) a ‘recognition loop’, which acts as a static anchor at the ETF–partner interface, and (b) a highly mobile redox‐active FAD domain. Together, this enables the FAD domain of ETF to sample a range of conformations, some compatible with fast interprotein electron transfer. This ‘conformational sampling’ enables ETF to recognize structurally distinct partners, whilst also maintaining a degree of specificity. Complex formation triggers mobility of the FAD domain, an ‘induced disorder’ mechanism contrasting with the more generally accepted models of protein–protein interaction by induced fit mechanisms. We discuss the implications of the highly dynamic nature of ETFs in biological interprotein electron transfer. ETF complexes point to mechanisms of electron transfer in which ‘dynamics drive function’, a feature that is probably widespread in biology given the modular assembly and flexible nature of biological electron transfer systems.

The field of synthetic biology aims to make the design of biological systems predictable, shrinking the huge design space to practical numbers for testing. When designing microbial cell factories, most optimisation efforts have focussed on enzyme and strain selection/engineering, pathway regulation and process development. In silico tools for the predictive design of bacterial ribosome binding sites (RBSs) and RBS libraries now allow translational tuning of biochemical pathways; however, methods for predicting optimal RBS combinations in multigene pathways do not currently exist. Here we present the implementation of machine learning algorithms to model the RBS sequence–phenotype relationship from representative subsets of large combinatorial RBS libraries allowing the accurate prediction of optimal high-producers. Applied to a recombinant monoterpenoid production pathway in Escherichia coli our approach was able to boost production titres by over 60 % when screening under 3 % of a library. To facilitate library screening, a multi-well plate fermentation procedure was developed allowing increased screening throughput with sufficient resolution to discriminate between high and low producers. High producers from one library did not translate during scale-up, but the reduced screening requirements allowed rapid re-screening at the larger scale. This methodology is potentially compatible with any biochemical pathway and provides a powerful tool towards predictive design of bacterial production chassis.

The crystal structure of the human electron transferring flavoprotein (ETF)·medium chain acyl-CoA dehydrogenase (MCAD) complex reveals a dual mode of protein-protein interaction, imparting both specificity and promiscuity in the interaction of ETF with a range of structurally distinct primary dehydrogenases. ETF partitions the functions of partner binding and electron transfer between (i) the recognition loop, which acts as a static anchor at the ETF·MCAD interface, and (ii) the highly mobile redox active FAD domain. Together, these enable the FAD domain of ETF to sample a range of conformations, some compatible with fast interprotein electron transfer. Disorders in amino acid or fatty acid catabolism can be attributed to mutations at the protein-protein interface. Crucially, complex formation triggers mobility of the FAD domain, an induced disorder that contrasts with general models of protein-protein interaction by induced fit mechanisms. The subsequent interfacial motion in the MCAD·ETF complex is the basis for the interaction of ETF with structurally diverse protein partners. Solution studies using ETF and MCAD with mutations at the protein-protein interface support this dynamic model and indicate ionic interactions between MCAD Glu²¹² and ETF Argα²⁴⁹ are likely to transiently stabilize productive conformations of the FAD domain leading to enhanced electron transfer rates between both partners.