A bifunctional asparaginyl endopeptidase efficiently catalyzes both cleavage and cyclization of cyclic trypsin inhibitors.

Asparaginyl endopeptidases (AEPs) catalyze the key backbone cyclization step during the biosynthesis of plant-derived cyclic peptides. Here, we report the identification of two AEPs from Momordica cochinchinensis and biochemically characterize MCoAEP2 that catalyzes the maturation of trypsin inhibitor cyclotides. Recombinantly produced MCoAEP2 catalyzes the backbone cyclization of a linear cyclotide precursor (MCoTI-II-NAL) with a kcat/Km of 620 mM-1 s-1, making it one of the fastest cyclases reported to date. We show that MCoAEP2 can mediate both the N-terminal excision and C-terminal cyclization of cyclotide precursors in vitro. The rate of cyclization/hydrolysis is primarily influenced by varying pH, which could potentially control the succession of AEP-mediated processing events in vivo. Furthermore, MCoAEP2 efficiently catalyzes the backbone cyclization of an engineered MCoTI-II analog with anti-angiogenic activity. MCoAEP2 provides enhanced synthetic access to structures previously inaccessible by direct chemistry approaches and enables the wider application of trypsin inhibitor cyclotides in biotechnology applications.

Creating Disease Resistant Chickens: A Viable Solution to Avian Influenza?

Influenza A virus (IAV) represents an ongoing threat to human and animal health worldwide. The generation of IAV-resistant chickens through genetic modification and/or selective breeding may help prevent viral spread. The feasibility of creating genetically modified birds has already been demonstrated with the insertion of transgenes that target IAV into the genomes of chickens. This approach has been met with some success in minimising the spread of IAV but has limitations in terms of its ability to prevent the emergence of disease. An alternate approach is the use of genetic engineering to improve host resistance by targeting the antiviral immune responses of poultry to IAV. Harnessing such resistance mechanisms in a "genetic restoration" approach may hold the greatest promise yet for generating disease resistant chickens. Continuing to identify genes associated with natural resistance in poultry provides the opportunity to identify new targets for genetic modification and/or selective breeding. However, as with any new technology, economic, societal, and legislative barriers will need to be overcome before we are likely to see commercialisation of genetically modified birds.