Institution: | Department of Biochemistry, Royal Holloway and Bedford New College, Egham Surrey, TW20 0EX, U.K. a Department of Biochemistry, University of the Basque Country, P.O. Box 644, Bilbao, Spain |
Abstract: | Strains of car B (phytoene-accumulating) mutants of Phycomyces blakesleeanus have been characterized with respect to their carotene contents, in vitro formation of isoprenoids from 2-14C] mevalonic acid and their ability to produce 14C]phytoene in situ for use in coupled assays of phytoene desaturase activity. All strains produced predominantly (15-Z)-phytoene both in vivo and in vitro. Other isoprenoids were produced by cell extracts including squalene, sterols, prenyl diphosphates and prenyl alcohols. The addition of 1% Tween 60 to crude cell extracts of the mutants partially restored wild type carotenogenic activity and also altered the proportions of other isoprenoids formed. However, in a cytosolic fraction of the car B mutant, the addition of 1% Tween 60 did not result in the production of any carotenoid from phytoene. This fraction was the most effective source of 14C] phytoene for use in coupled assays of phytoene desaturase activity. |