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Proteins, Expression, Isolation and Analysis> | … for the CCD of Chrysanthemum morifolium (CmCCD4, UniProtKB B0FLM7) and Crocus sativus (CsCCD4, UniProtKB B7SNW2), and the N-terminal region, corresponding to the first 210 amino acids, of VP14 (UniProtKB O24592) were synthesized by Genscript (Piscataway, NJ … | Get A Quote |
Synthesis of β-ionone in recombinant Saccharomyces cerevisiae is limited by the efficiency of Carotenoid Cleavage Dioxygenases (CCD), membrane-tethered enzymes catalyzing the last step in the pathway. We performed in silico design and membrane affinity analysis, focused on single-point mutations of PhCCD1 to improve membrane anchoring. The resulting constructs were tested in a β-carotene hyper-producing strain by comparing colony pigmentation against colonies transformed with native PhCCD1 and further analyzed by β-ionone quantification via RP-HPLC. Two single-point mutants increased β-ionone yields almost 3-fold when compared to native PhCCD1. We also aimed to improve substrate accessibility of PhCCD1 thro... More