miniinteins

Miniinteins are shortened versions of inteins in which the homing endonuclease domain is absent or greatly reduced. Like full-length inteins, mini-inteins undergo a self-splicing reaction that removes the intein from a host protein and joins the flanking extein segments. Removal of the endonuclease domain yields a smaller polypeptide, which often improves expression, solubility, and compatibility with fusion-tag strategies in recombinant protein production.

Most naturally occurring inteins contain an endonuclease domain that promotes mobility; miniinteins retain the core splicing machinery but lack this domain, reducing potential unwanted nuclease activity. This reduction can also simplify genetic constructs and streamline engineering workflows.

Common uses of miniinteins include self-cleaving affinity tags and native terminus generation in recombinant proteins. The IMPACT technology, which uses the Mxe GyrA mini-intein, can produce proteins with native N- or C-termini after cleavage. Mini-inteins are also employed in protein ligation and cyclization through split-intein approaches, enabling controlled assembly of polypeptides under mild conditions.

Important considerations for miniinteins include extein sequence constraints at the splice junctions, which can affect splicing efficiency, and variable kinetics across different miniintein variants and host environments. Some mini-inteins exhibit robust performance in a range of conditions, while others require optimization of junction residues or reaction parameters.

Ongoing research aims to expand the repertoire of miniinteins with greater tolerance to extein sequences, faster splicing, and compatibility with diverse organisms. The development of orthogonal mini-intein systems also seeks to enable complex multi-step protein engineering in single or multi-host contexts.