Preparing GST-, His-, or MBP-Fusion Proteins from Bacteria

An excellent source of antigens is to overexpress cloned genes in bacteria. A wide variety of coding regions can be expressed in bacteria either on their own or as fusion proteins. Indeed, it is often convenient to use vectors that express the antigen fused to an affinity tag, such as glutathione-S-transferase (GST), maltose-binding protein (MBP), or poly-histidine (e.g., 6×His). Each of these tags reliably binds to a specific affinity column enabling the antigen to be conveniently eluted under appropriate conditions. It should be noted that GST adds ∼25 kDa and MBP adds 40 kDa to the expressed protein. Because bacteria will reliably express proteins that are <80 kDa, the mass of the affinity tag should be included when designing your recombinant protein construct.