Consecutive interactions of Lep H2 during membrane insertion. (A) Schematic representation of the 72–222Lep constructs with a C-terminal Myc tag and the crosslinking probe at position 71 or 73. (B) 102–222LepTAG71 and 73 were produced, crosslinked, carbonate extracted and immunoprecipitated as described in Fig 2. (C) Quantification of nascent Lep photo-crosslinked to YidC relative to the amount of immunoprecipitated nascent chains using the anti-Myc serum. The highest value for crosslinking efficiency was taken as 100%. (D) To identify lipid crosslinking, 102–162LepTAG73 constructs were produced, crosslinked or kept in the dark, immunoprecipitated using anti-Myc serum and analysed by 16% tricine SDS–PAGE (indicated by asterisks).