Fig. 2
SDS-PAGE and HPLC separation of HisrMlat1 from M15 strain cells. a SDS-PAGE. Left lane shows the molecular weight markers (kDa). Lane 1: the cell protein content that has the pQE30 vector without the HisrMlat1 gene and without IPTG induction. Lane 2: the cell protein content that has the pQE30 vector without the HisrMlat1 gene after 0.1 mM IPTG induction. Lane 3: the cell protein content that has the pQE30 vector with the HisrMlat1 gene and without IPTG induction. Lane 4: the cell protein content that has the pQE30 vector with the HisrMlat1 gene and with 0.1 mM IPTG induction. Lane 5: the protein after purification by affinity column (upper bands are oligomers of HisrMlat1). The protein band corresponding to HisrMlat1 is indicated by an arrow. b rpHPLC. The product obtained from the affinity column was directly loaded into the C18 reverse column (approx. 100 μg of protein). The fractions labeled with an asterisk had the molecular mass expected for the recombinant HisrMlat1 (label i). None of the fractions were lethal to mice. The pooled multiple oxidized fractions (100 μg of protein, label i) were reduced with DTT, and loaded into the same rpHPLC. Superimposed on this chromatogram is the profile of the reduced HisrMlat1 (label ii)