title: Modulation of intein activity by its neighboring extein substrates.
abstract: Inteins comprise a large family of phylogenetically widespread self-splicing protein catalysts that colonize diverse host proteins. The evolutionary and functional relationship between the intein and the split-host protein, the exteins, is largely unknown. To probe an association, we developed an in vivo and in vitro intein assay based on FRET. The FRET assay reports cleavage of the intein from its N-terminal extein. Applying this assay to randomized extein libraries, we show that the nature of the extein substrate bordering the intein can profoundly influence intein activity. Residues proximal to the intein-splicing junction in both N- and C-terminal exteins can accelerate the N-terminal cleavage rate by >4-fold or attenuate cleavage by 1,000-fold, both resulting in compromised self-splicing efficiency. The existence and the magnitude of extein effects require consideration for maximizing the utility of inteins in biotechnological applications, and they predict biases in intein integration sites in nature.
authors.affiliation: Center for Medical Science, New York State Department of Health, 150 New Scotland Avenue, Albany, NY 12208, USA.
authors.forename: Gil
authors.initials: G
authors.lastname: Amitai
authors.forename: Brian P
authors.initials: BP
authors.lastname: Callahan
authors.forename: Matt J
authors.initials: MJ
authors.lastname: Stanger
authors.forename: Georges
authors.initials: G
authors.lastname: Belfort
authors.forename: Marlene
authors.initials: M
authors.lastname: Belfort
mesh_terms: D000596:Amino Acids / Q000378:metabolism; D047669:Exteins; D031541:Fluorescence Resonance Energy Transfer; D047668:Inteins; D050505:Mutant Proteins / Q000378:metabolism; D019154:Protein Splicing
chemical_list: D000596:Amino Acids; D050505:Mutant Proteins