Fig. 2: Protein trans-splicing of the wildtype Aes intein. | Nature Communications

Fig. 2: Protein trans-splicing of the wildtype Aes intein.

From: A cysteine-less and ultra-fast split intein rationally engineered from being aggregation-prone to highly efficient in protein trans-splicing

Fig. 2

a Scheme of the PTS reaction in a simplified three-state kinetic model. The AesN and AesC fragments are 120 and 39 aa in size, respectively. Not shown are the linear intermediate resulting from the N-O acyl shift and the C-terminal succinimide prior to hydrolysis. b SDS-PAGE analysis of the PTS reaction at 37 °C using precursors 1P and 2P at 10 µM and 30 µM, respectively, and in the absence of any reducing agents. This experiment was repeated three times. c Time-course of the PTS reaction obtained by densitometric analysis of data as shown in (b) and three-state kinetic model fitted to the data. d Splice product formation plotted against the time applying different molar ratios of the precursors 1P and 3P. Calculated molecular masses are: PN(1P) = 58.2 kDa, PC(2P) = 31.3 kDa, BI = 74.8 kDa, SP = 70.1 kDa, AesN = 14.7 kDa, AesC = 4.7 kDa. BI branched intermediate, SP splice product. For (c, d), n = 3 technical replicates. Data are presented as mean ± s.d. normalized to the molecular weight of each protein species. Source data are provided as a Source Data file.

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