An intein from Halobacterium salinarum can be isolated as an unspliced precursor protein with exogenous exteins after Escherichia coli overexpression. The intein promotes protein splicing and uncoupled N-terminal cleavage in vitro, conditional on incubation with NaCl or KCl at concentrations of >1.5 M. The protein splicing reaction also is conditional on reduction of a disulfide bond between two active site cysteines. Conditional protein splicing under these relatively mild conditions may lead to advances in intein-based biotechnology applications and hints at the possibility that this H. salinarum intein could serve as a switch to control extein activity under physiologically relevant conditions.

Protein splicing is a post‐translational modification where an internal polypeptide, called an intein, catalyzes its excision from two flanking polypeptides, called exteins, concomitant with extein ligation. Usually, when intein fusion proteins from other organisms are overexpressed in E. coli, splicing is spontaneous in vivo. However, despite a very similar sequence to the Mma PolII intein, which is capable of splicing, the Halobacterium salinarum (Hsa) PolII intein did not splice upon in vivo expression in E. coli. Halobacterium salinarum is a halophilic organism, so it lives in an environment with extremely high salt concentrations. In an effort to induce reactivity, two fusion proteins of the Hsa intein were incubated at 28°C for 16 hours with increasing amounts of NaCl. The fusion protein with a wild type intein has glutamine as the last residue of the intein, whereas the second intein, termed MIHQN, replaces this glutamine with asparagine. After incubation, in vitro splicing was observed in both intein fusion proteins on incubation with 2 M and 2.5 M NaCl.Support or Funding InformationThis material is based upon work supported by the National Science Foundation under grants MCB‐1244089 and MCB‐1517138 (KVM), by a Henry Dreyfus Teacher‐Scholar Award (KVM), and the Summer Research Fellowships from Renee and Anthony M. Marlon, M.D (CC).