title: Rational reprogramming of fungal polyketide first-ring cyclization.
abstract: Resorcylic acid lactones and dihydroxyphenylacetic acid lactones represent important pharmacophores with heat shock response and immune system modulatory activities. The biosynthesis of these fungal polyketides involves a pair of collaborating iterative polyketide synthases (iPKSs): a highly reducing iPKS with product that is further elaborated by a nonreducing iPKS (nrPKS) to yield a 1,3-benzenediol moiety bridged by a macrolactone. Biosynthesis of unreduced polyketides requires the sequestration and programmed cyclization of highly reactive poly-β-ketoacyl intermediates to channel these uncommitted, pluripotent substrates to defined subsets of the polyketide structural space. Catalyzed by product template (PT) domains of the fungal nrPKSs and discrete aromatase/cyclase enzymes in bacteria, regiospecific first-ring aldol cyclizations result in characteristically different polyketide folding modes. However, a few fungal polyketides, including the dihydroxyphenylacetic acid lactone dehydrocurvularin, derive from a folding event that is analogous to the bacterial folding mode. The structural basis of such a drastic difference in the way a PT domain acts has not been investigated until now. We report here that the fungal vs. bacterial folding mode difference is portable on creating hybrid enzymes, and we structurally characterize the resulting unnatural products. Using structure-guided active site engineering, we unravel structural contributions to regiospecific aldol condensations and show that reshaping the cyclization chamber of a PT domain by only three selected point mutations is sufficient to reprogram the dehydrocurvularin nrPKS to produce polyketides with a fungal fold. Such rational control of first-ring cyclizations will facilitate efforts to the engineered biosynthesis of novel chemical diversity from natural unreduced polyketides.
authors.affiliation: Natural Products Center, School of Natural Resources and the Environment, University of Arizona, Tucson, AZ 85706, USA.
authors.forename: Yuquan
authors.initials: Y
authors.lastname: Xu
authors.forename: Tong
authors.initials: T
authors.lastname: Zhou
authors.forename: Zhengfu
authors.initials: Z
authors.lastname: Zhou
authors.forename: Shiyou
authors.initials: S
authors.lastname: Su
authors.forename: Sue A
authors.initials: SA
authors.lastname: Roberts
authors.forename: William R
authors.initials: WR
authors.lastname: Montfort
authors.forename: Jia
authors.initials: J
authors.lastname: Zeng
authors.forename: Ming
authors.initials: M
authors.lastname: Chen
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.forename: Min
authors.initials: M
authors.lastname: Lin
authors.forename: Jixun
authors.initials: J
authors.lastname: Zhan
authors.forename: István
authors.initials: I
authors.lastname: Molnár
mesh_terms: D000447:Aldehydes / Q000737:chemistry; D001483:Base Sequence; D053898:Biosynthetic Pathways / Q000502:physiology*; D020134:Catalytic Domain / Q000235:genetics; D003001:Cloning, Molecular; D003500:Cyclization / Q000502:physiology; D004926:Escherichia coli; D005285:Fermentation; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D048630:Polyketide Synthases / Q000378:metabolism*; D061065:Polyketides / Q000378:metabolism*; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis*; D017422:Sequence Analysis, DNA
chemical_list: D000447:Aldehydes; D061065:Polyketides; D029701:Saccharomyces cerevisiae Proteins; D048630:Polyketide Synthases; C116609:3-hydroxybutanal