Project: Whole-cell Mediated, Stereoselective Oxidation of Organic Compounds Using Bacterial Environmental Isolates

Biocatalytic reactions provide many of the chiral building blocks needed for drug synthesis and microbial degradation is one of the few practical methods for large-scale bioremediation of persistent organic pollutants. Available commercial enzymes and microorganisms often fail to convert specific substrates to desired products, creating an ongoing need to find new organisms. Environmental samples are being screened to obtain new microbes for whole-cell mediated preparation of chiral sulfoxides (Scheme 1), among other targets.

Scheme 1.

Chiral oxidation of sulfides using biocatalysis is the simplest and most economical route to these versatile synthetic intermediates, but studies using known monooxygenases and dioxygenases from deposited cultures showed that substrate ranges and enantioselectivities for these systems are limited. Our primary collection site (Sulphur Mountain, Ventura County, CA) is a geologic anomaly rich in both sulfur and petroleum hydrocarbons, which bacteria likely are able to detoxify and/or utilize. Already, students have isolated and are characterizing over one hundred bacterial monocultures from soil and water samples, and currently are monitoring chemical transformations induced by these microbes in terms of a library of substrates, which include prochiral sulfides, dibenzothiophene, toluene, and pyridine. Our pure strains are compatible with and/or consume the generally-toxic thioanisole (Scheme 1, prochiral sulfide) and have been identified by 16S rRNA gene sequence analysis as Pseudomonas spp.

The next phase of this research will involve the following objectives:

·         Rapid screening of a library of 25 strains to identify which are most able to tolerate/utilize the above carbon sources,

·         Grow the best strains on a larger scale in the presence of the organic substrate and follow the conversion by GC/MS,

·         Isolate the main organic products and fully characterize them,

·         Analyze enantiomeric excesses (e.e.’s) in the products, where relevant,

·         Investigate the impact of the culturing conditions on product yield and e.e.’s.

Experience in microbiology, particularly aseptic technique, and organic chemistry (theory and lab) will be considered as a plus.