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. Author manuscript; available in PMC: 2020 May 3.
Published in final edited form as: J Mol Biol. 2019 Mar 23;431(10):2006–2019. doi: 10.1016/j.jmb.2019.03.021

Figure 2.

Figure 2.

To be recognized by SecA, the 16 residue Ala/Leu H-segment must have 3–4 leucines in order to be recognized by SecA. This immunoblot identifies T7 tags, which are contained within the N-terminal domain carrying the H-segment. The appearance of the lower molecular weight bands indicates cleavage by SPase I and therefore processing of the construct by SecA. The seven H-segments used in the constructs are indicated. The blots show that to be recognized by SecA, the H-segment must contain at least 3 leucines (lane H3), but for complete processing 4 leucines are required (lane H4). A single Leu-to-Pro substitution at the center of the A13L6 construct to give A13P1L5 prevents recognition and secretion of by SecA. The C-H-F constructs were expressed in E. coli BL21 cells grown in SOC media at 37° C (see Materials and Methods).