Nucleic Acids Res. doi:10.1093/nar/gkv612

Inteins are self-splicing elements that excise themselves from translated proteins to leave behind the extein, or mature protein. The majority of functionally relevant residues are thought to be internal to the intein, though a few residues in the extein sequence directly bordering the intein have been shown to influence function. Some inteins are known to be conditionally regulated, but most documented cases have focused on intein-specific mechanisms, and it is not known whether exteins could have a larger role in regulating splicing. To investigate this question, Topilina et al. considered the RadA/RecA family of recombinases, in which the intein is inserted in one of five points—two newly reported in this work—either at monomer-monomer interfaces or within the ATP-binding site. Comparison of the intein from the hyperthermophile Pyrococcus horikoshii RadA in its native site and within an engineered MBP-GFP construct demonstrated that the MBP-GFP product formed very efficiently, whereas the native construct showed no splicing at 25 °C even after several days. Splicing activity could be increased either by raising the temperature or by adding an ionic liquid and detergent; splicing was also coincident with changes in protein structure, as monitored by CD, but not in ATPase activity, suggesting that the flexibility of the extein was moderating intein splicing. A model of the unspliced RadA pointed to several charged residues in the extein in close proximity physically, but not sequentially, to functionally important intein residues; mutation of these residues, but not charged residues elsewhere in the sequence, led to substantial increases in splicing. These combined results led to a model in which the extein-intein interaction at low temperatures prevents intein splicing, while temperatures akin to those in the organism's native environment release these contacts to promote splicing and RadA activation. The authors speculate that this regulation may help prevent ATP consumption under stressful conditions.