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. 2022 Aug 12;62(2):292–299. doi: 10.1021/acs.biochem.2c00241

Figure 2.

Figure 2

Polyhistidine length impacts purification of spike-His-Fer, but not HA-His-Fer. (A) Site 4, as in Figure 1, highlighting the length of the installed polyhistidine tag, ranging from four to eight histidine residues. The image in panel A shows installation of six histidine residues. Dot blots tracking the purification of His-4.4–His-4.8 proteins for either (B) HA or (C) spike. Columns show dots from the load, flow through (FT), wash (W), or elution (Elu). Expression was detected for all lengths of HA-His-Fer but only for greater than or equal to six histidine residues for spike-His-Fer. (D) SDS–PAGE analysis of the elution from the dot blots shown in panels B and C. Lanes in HA-His-Fer correspond to a molecular weight (MW), in kilodaltons, ladder (lane 1, labels on the left), His-4.4–His-4.8 (lanes 2–6, respectively), and the WT (lane 7). Lanes in spike-His-Fer correspond to the MW ladder (lane 1, labels on the left), the WT (lane 2), and His-4.4–His-4.8 (lanes 3–7, respectively). These results confirm the dot blot analysis and demonstrate that the proteins migrate at the expected MW.