FIGURE 6.
Carbonylation of HbpS amino acid side-chains. Untreated (lane 1) or treated with FeCl2 (100 μm, lane 2; 250 μm, lane 3; 500 μm, lane 4) and 5 mm DTT HbpS WT proteins were subjected to DNP derivatization as described under “Materials and Methods.” Reaction products were loaded, without previous boiling, onto 12% SDS-polyacrylamide gels. After electrophoresis, proteins were analyzed by Coomassie staining (A) or by detecting derivatized DNP groups by immunoblotting using anti-DNP antibodies (B). Arrows indicate positions for the monomeric (1mer), dimeric (2mer), or tetrameric (4mer) HbpS forms. The approximate sizes of prestained protein markers are also shown (lane 5).