Development of a whole-cell biosensor for ethylene oxide and ethylene.

Ethylene and ethylene oxide are widely used in the chemical industry, and ethylene is also important for its role in fruit ripening. Better sensing systems would assist risk management of these chemicals. Here, we characterise the ethylene regulatory system in Mycobacterium strain NBB4 and use these genetic parts to create a biosensor. The regulatory genes etnR1 and etnR2 and cognate promoter Petn were combined with a fluorescent reporter gene (fuGFP) in a Mycobacterium shuttle vector to create plasmid pUS301-EtnR12P. Cultures of M. smegmatis mc2-155(pUS301-EtnR12P) gave a fluorescent signal in response to ethylene oxide with a detection limit of 0.2 µM (9 ppb). By combining the epoxide biosensor cells with another culture expressing the ethylene monooxygenase, the system was converted into an ethylene biosensor. The co-culture was capable of detecting ethylene emission from banana fruit. These are the first examples of whole-cell biosensors for epoxides or aliphatic alkenes. This work also resolves long-standing questions concerning the regulation of ethylene catabolism in bacteria.

A novel soluble di-iron monooxygenase from the soil bacterium Solimonas soli.

Soluble di-iron monooxygenase (SDIMO) enzymes enable insertion of oxygen into diverse substrates and play significant roles in biogeochemistry, bioremediation and biocatalysis. An unusual SDIMO was detected in an earlier study in the genome of the soil organism Solimonas soli, but was not characterized. Here, we show that the S. soli SDIMO is part of a new clade, which we define as 'Group 7'; these share a conserved gene organization with alkene monooxygenases but have only low amino acid identity. The S. soli genes (named zmoABCD) could be functionally expressed in Pseudomonas putida KT2440 but not in Escherichia coli TOP10. The recombinants made epoxides from C2 C8 alkenes, preferring small linear alkenes (e.g. propene), but also epoxidating branched, carboxylated and chlorinated substrates. Enzymatic epoxidation of acrylic acid was observed for the first time. ZmoABCD oxidised the organochlorine pollutants vinyl chloride (VC) and cis-1,2-dichloroethene (cDCE), with the release of inorganic chloride from VC but not cDCE. The original host bacterium S. soli could not grow on any alkenes tested but grew well on phenol and n-octane. Further work is needed to link ZmoABCD and the other Group 7 SDIMOs to specific physiological and ecological roles.