Forskolin is a unique structurally complex labdane-type diterpenoid used in the treatment of glaucoma and heart failure based on its activity as a cyclic AMP booster. Commercial production of forskolin relies exclusively on extraction from its only known natural source, the plant Coleus forskohlii, in which forskolin accumulates in the root cork. Here, we report the discovery of five cytochrome P450s and two acetyltransferases which catalyze a cascade of reactions converting the forskolin precursor 13R-manoyl oxide into forskolin and a diverse array of additional labdane-type diterpenoids. A minimal set of three P450s in combination with a single acetyl transferase was identified that catalyzes the conversion of 13R-manoyl oxide into forskolin as demonstrated by transient expression in Nicotiana benthamiana. The entire pathway for forskolin production from glucose encompassing expression of nine genes was stably integrated into Saccharomyces cerevisiae and afforded forskolin titers of 40 mg/L.DOI: http://dx.doi.org/10.7554/eLife.23001.001

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Enzymatic protein hydrolysis (EPH) is one of the industrial bioprocesses used to recover valuable constituents from food processing by-products. Extensive heterogeneity of byproducts from, for example, meat-processing is a major challenge in production of protein hydrolysates with stable and desirable quality attributes. Therefore, there is a need for process control tools for production of hydrolysates with defined qualities from such heterogeneous raw materials. In the present study, we are reporting a new feed-forward process control strategy for enzymatic protein hydrolysis of poultry by-products. Four different spectroscopic techniques, i.e., NIR imaging scanner, a miniature NIR (microNIR) instrument, fluorescence and Raman, were evaluated as tools for characterization of the raw material composition. Partial least squares (PLS) models for ash, protein and fat content were developed based on Raman, fluorescence and microNIR measurements, respectively. In an effort to establish feed-forward process control tools, we developed statistical models that enabled prediction of end-product characteristics, i.e., protein yield and average molecular weight of peptides (Mw), as a function of raw material quality and hydrolysis time. A multi-block sequential orthogonalised-PLS (SO-PLS) model, where spectra from one or more techniques and hydrolysis time were used as predictor variables, was fitted for the feed-forward prediction of product qualities. The best model was obtained for protein yield based on combined use of microNIR and fluorescence (R 2 = 0.88 and RMSE = 4.8). A Raman-based model gave a relatively moderate prediction model for Mw (R 2 = 0.56 and RMSE = 150). Such statistical models based on spectroscopic measurements of the raw material can be vital process control tools for EPH. To our knowledge, the present work is the first example of a spectroscopic feed-forward process control for an industrially relevant bioprocess.

A low molecular weight peptide fraction from chicken by-product hydrolysate was found to inhibit DPP-IV in vitro and promote cellular glucose uptake ex vivo.