Conditional protein splicing

The discovery of split inteins further advanced the scope of EPL into the realm of conditional protein splicing. As outlined above, certain inteins can be split into two halves. An advantage for a general applicability concerns the observation that the split site is asymmetric. This opens an access to chemical synthesis of the smaller intein half and hence an easy route to C-terminal modification of proteins with, e.g., fluorophores, lipid anchor, or other posttranslational modifications of proteins. 

An intriguing new development relates to conditional protein splicing [27]. In this scenario the property of the immunosuppressant rapamycin is utilized to bind simultaneously to two different proteins (FKBP and FRB) with high affinity. If FKBP and FRB are linked to the N- and C-termini of the intein-extein halves (see Fig. 4 for the protein constructs) the addition of rapamycin would trigger the formation of a ternary complex, thereby bringing the intein halves into close proximity. In a consecutive reaction the two exteins can combine to generate an active or, e.g., fluorescent labeled protein. Since both halves can be expressed in mammalian and other cells, cellular functions can be studied. 

Mootz HD, Blum ES, Tyszkiewicz AB, Muir TW (2003) Conditional protein splicing a new tool to control protein structure and function in vitro and in vivo. J Am Chem Soc 125 10561-10569... 

H.D. Mootz, E.S. Blum, T.W. Muir, Activation of an autoregulated protein kinase by conditional protein splicing,... 

A subsequent study in 2002 of 27 families with a condition known as multiminicore disease (MmD) also linked mutations in SEPNl to disease pathology. Multiple mutations were identified in exons 1, 5, 7, 8, 10, and 11, and the authors also mentioned that this region (RSMD) had been previously linked to MmD. Minicores are lesions by histochemistry of mitochondrial depletion within muscle tissue. The first biochemical study of selenoprotein N aimed to identify the protein localization by immunohistochemistry and found that the primary protein product of several identified mRNAs (splice variants) was a 70 kDa protein present in the endoplasmic reticulum. Two potential ER targeting domains were shown to be present and the peptide expressed from the first exon was shown to be required for localization into the ER. This study also revealed that selenoprotein N was an integral membrane protein that is N-glycosylated. Expression analysis showed pronounced levels in embryonic tissue with a reduction after development and differentiation. 

These are all the possibilities, beginning with exon-1. If we relax that condition, then we can clearly make several more exon-2-exon-3, for example. These proteins are known as splice variants. It follows that a genome coding for 25,000 proteins can actually generate quite a few more than that. Complexity gets stacked on complexity. 

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