Identification of a Possible Selenite Sensor Protein from Enterobacter sp. YSU

Abstract

The Y-12 plant (Oak Ridge, TN) contributed to national defense during World War II and the cold war. The plant processed uranium to make nuclear bombs and later switched to lithium processing to make hydrogen bombs. This process resulted in heavy metal waste deposits in East Fork Poplar Creek and the surrounding environment. Enterobacter sp. YSU, which was isolated from this creek, was found to be resistant to metal salts of zinc, cadmium, mercury, selenite, silver, copper and gold. Metal resistant bacteria encode proteins involved in pumping metals out of the cell, in converting them to less toxic forms or in sequestering them. Transposon mutagenesis was used to identify genes involved in selenite resistance. When introduced into the YSU strain, the EZ-Tn5TM<R6Kγ ori/KAN-2> Tnp TransposomeTM inserted itself randomly into the genome of this bacterium. One of the transposome transformants, L31, was sensitive to selenite on agar plates. Minimal inhibitory concentration (MIC) experiments in liquid cultures showed that it was sensitive to 40 mM selenite in LB medium, but not in M-9 minimal medium. Gene rescue, DNA sequencing and Basic Local Alignment Search Tool (BLAST) analysis showed that the interrupted gene product is related to a histidine kinase sensor protein. It may regulate a surrounding gene that encodes a protein involved in efflux of toxic metals.