Characterisation of a novel soluble di-iron monooxygenase from the soil organism Solimonas soli

Abstract

Monooxygenase enzymes are responsible for the oxidation of hydrocarbons and other compounds in the carbon and nitrogen cycles, are important for the biodegradation of pollutants, and can act as biocatalysts for chemical manufacturing. The soluble di-iron monooxygenases (SDIMOs) are of interest due to their broad substrate range, high enantioselectivity, and ability to oxidise inert substrates like methane. An unusual SDIMO was detected in an earlier study in the genome of the soil organism Solimonas soli but was not characterised. This study has shown that the S. soli SDIMO is part of a new SDIMO clade, which is defined as ‘Group 7’. The S. soli group 7 SDIMO genes (named zmoABCD) was functionally expressed in Pseudomonas putida KT2440 and the recombinants made epoxides from C2-C8 alkenes, preferring small linear alkenes (especially propene). ZmoABCD also oxidised vinyl chloride (VC) and cis-1,2-dichloroethene (cDCE). However, the original host bacterium S. soli could not grow on any alkenes tested but grew well on phenol, noctane, isoleucine, leucine, and Tween 80. ΔzmoABCD knockout strains of S. soli were also able to grow using these substrates as sole carbon sources, suggesting that ZmoABCD is not the sole locus responsible for growth on these molecules. The regulation of zmo expression was also studied using a plasmid-borne bioreporter construct, where a gfp reporter gene was placed under the control of the native zmo promoter to identify potential inducers of zmoABCD. It was demonstrated in S. soli that Tween 80, butane, 1-butene, and 2-propanol induced the expression of zmoABCD while n-octane and 1-octene repressed expression. This study has provided a substantial framework to narrow down the substrate range of ZmoABCD. The characterisation of ZmoABCD will increase our understanding of SDIMO evolutionary history and enable new applications of these enzymes for biocatalysis and bioremediation.