Back to Search
Start Over
Engineered biosynthesis of plant polyketides: structure-based and precursor-directed approach.
- Source :
-
Topics in current chemistry [Top Curr Chem] 2010; Vol. 297, pp. 45-66. - Publication Year :
- 2010
-
Abstract
- Pentaketide chromone synthase (PCS) and octaketide synthase (OKS) are novel plant-specific type III polyketide synthases (PKSs) obtained from Aloe arborescens. Recombinant PCS expressed in Escherichia coli catalyzes iterative condensations of five molecules of malonyl-CoA to produce a pentaketide 5,7-dihydroxy-2-methylchromone, while recombinant OKS carries out sequential condensations of eight molecules of malonyl-CoA to yield octaketides SEK4 and SEK4b, the longest polyketides produced by the structurally simple type III PKS. The amino acid sequences of PCS and OKS are 91% identical, sharing 50-60% identity with those of other chalcone synthase (CHS) superfamily type III PKSs of plant origin. One of the most characteristic features is that the conserved active-site Thr197 of CHS (numbering in Medicago sativa CHS) is uniquely replaced with Met207 in PCS and with Gly207 in OKS, respectively. Site-directed mutagenesis and X-ray crystallographic analyses demonstrated that the chemically inert single residue lining the active-site cavity controls the polyketide chain length and the product specificity depending on the steric bulk of the side chain. On the basis of the crystal structures, an F80A/Y82A/M207G triple mutant of the pentaketide-producing PCS was constructed and shown to catalyze condensations of nine molecules of malonyl-CoA to produce an unnatural novel nonaketide naphthopyrone, whereas an N222G mutant of the octaketides-producing OKS yielded a decaketide benzophenone SEK15 from ten molecules of malonyl-CoA. On the other hand, the type III PKSs exhibited broad substrate specificities and catalytic potential. OKS accepted p-coumaroyl-CoA as a starter substrate to produce an unnatural novel C19 hexaketide stilbene and a C21 heptaketide chalcone. Remarkably, the C21 chalcone-forming activity was dramatically increased in the structure-guided OKS N222G mutant. In addition, OKS N222G mutant also yielded unnatural novel polyketides from phenylacetyl-CoA and benzoyl-CoA as a starter substrate. These results suggested that the engineered biosynthesis of plant polyketides by combination of the structure-based and the precursor-directed approach would lead to further production of chemically and structurally divergent unnatural novel polyketides.
- Subjects :
- Amino Acid Sequence
Crystallography, X-Ray
Models, Molecular
Molecular Sequence Data
Mutagenesis, Site-Directed
Plants chemistry
Plants genetics
Polyketide Synthases chemistry
Protein Engineering trends
Sequence Alignment
Substrate Specificity
Macrolides chemistry
Macrolides metabolism
Plants enzymology
Polyketide Synthases genetics
Polyketide Synthases metabolism
Protein Engineering methods
Subjects
Details
- Language :
- English
- ISSN :
- 0340-1022
- Volume :
- 297
- Database :
- MEDLINE
- Journal :
- Topics in current chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 21495256
- Full Text :
- https://doi.org/10.1007/128_2009_22