Proteomic Analysis of MTN Deficiency in Enterohemorrhagic E. coli O157:H7
Faculty Mentor Information
Dr. Ken Cornell and Dr. Shin Pu
Abstract
The bacterial enzyme 5’-methylthioadenosine/S-adenosylhomocysteine nucleosidase (MTN) is essential for the production of autoinducers, and required to salvage methionine and purine constituents from S-adenosyl-methionine (SAM) dependent reactions. MTN represents a potentially valuable target for the development of novel antibiotics that would attenuate virulence by quorum sensing. To further study the impact of MTN activity on cellular processes, the proteomic profiles of enterohemorrhagic E. coli (EHEC) strain O157:H7 wild type (WT) and MTN knock-out (KO) cells were analyzed using shot-gun proteomic approach following peptide labeling and using liquid chromatography-tandem mass spectrometry. The results indicate that loss of MTN activity causes numerous changes in the expression of metabolic proteins. The MTN KO strain showed altered expression levels of enzymes responsible for methionine biosynthesis,spermidine biosynthesis, and radical SAM reactions that lead to the synthesis of vitamins (thiamine, lipoate, biotin). The decrease in vitamin synthesis may explain the decreases in activity of pathways involved in energy metabolism in the KO strain. Ultimately, the results suggest that antibiotics targeting MTN activity may function by widespread metabolic interruption. The impact of an MTN KO on the regulation of SAM-dependent methyltransferases, additional radical SAM enzyme activity, and polyamine dependent cellular activities will be explored in future research.
Proteomic Analysis of MTN Deficiency in Enterohemorrhagic E. coli O157:H7
The bacterial enzyme 5’-methylthioadenosine/S-adenosylhomocysteine nucleosidase (MTN) is essential for the production of autoinducers, and required to salvage methionine and purine constituents from S-adenosyl-methionine (SAM) dependent reactions. MTN represents a potentially valuable target for the development of novel antibiotics that would attenuate virulence by quorum sensing. To further study the impact of MTN activity on cellular processes, the proteomic profiles of enterohemorrhagic E. coli (EHEC) strain O157:H7 wild type (WT) and MTN knock-out (KO) cells were analyzed using shot-gun proteomic approach following peptide labeling and using liquid chromatography-tandem mass spectrometry. The results indicate that loss of MTN activity causes numerous changes in the expression of metabolic proteins. The MTN KO strain showed altered expression levels of enzymes responsible for methionine biosynthesis,spermidine biosynthesis, and radical SAM reactions that lead to the synthesis of vitamins (thiamine, lipoate, biotin). The decrease in vitamin synthesis may explain the decreases in activity of pathways involved in energy metabolism in the KO strain. Ultimately, the results suggest that antibiotics targeting MTN activity may function by widespread metabolic interruption. The impact of an MTN KO on the regulation of SAM-dependent methyltransferases, additional radical SAM enzyme activity, and polyamine dependent cellular activities will be explored in future research.