Say I have synthesized a new compound that gives great results against e. coli, how to know the best enzyme to target in e. coli? or how to predict the most possible target?
If the question is how to figure out the target of the compound with antibacterial activity, a common approach is to select for resistance by gradually increasing the concentration of the drug to a culture, then sequencing the genomes of the parental and resistant strains to see what has changed. This does not always work, since the resistance may be due to changes in efflux pumps or other changes resulting in non-specific resistance, but it is worth trying.
Another method is bacterial cytological profiling. This method can classify the pathway in which an antibacterial compound acts by its effect on the way certain dyes stain the bacteria.
This assumes that there is a specific binding target of this compound. You should also consider the possibility that this compound inhibits the bacteria by a non-specific mechanism, such as membrane disruption.
I suggest you watch the bacteria under a microscope when you apply the compound to see whether there are any obvious morphological changes that would provide a clue to the mechanism of action. For example, inhibitors of some processes result in prevention of cell separation after growth, leading to longer-than-normal bacteria. Membrane blebbing would be an indication that the compound acts on the cell membrane.
One alternative is to make photocrosslinkable analogs of the compound with a motif for click chemistry. You then can use UV to covalently bind the analogs to the targets and then use click chemistry to add groups for enrichment using coated beads. The pulled down proteins can then be identified using mass spectroscopy.
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