NDLI: Molecular Cloning, Modeling, and Site-Directed Mutagenesis of Type III Polyketide Synthase from Sargassum binderi (Phaeophyta)

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Molecular Cloning, Modeling, and Site-Directed Mutagenesis of Type III Polyketide Synthase from Sargassum binderi (Phaeophyta)

Content Provider	Springer Nature Link
Author	Baharum, Hariyanti Morita, Hiroyuki Tomitsuka, Akifumi Lee, Fong Chin Ng, Kim Yong Abdul Rahim, Raha Abe, Ikuro Ho, Chai Ling
Copyright Year	2010
Abstract	Type III polyketide synthases (PKSs) produce an array of metabolites with diverse functions. In this study, we have cloned the complete reading frame encoding type III PKS (SbPKS) from a brown seaweed, Sargassum binderi, and characterized the activity of its recombinant protein biochemically. The deduced amino acid sequence of SbPKS is 414 residues in length, sharing a higher sequence similarity with bacterial PKSs (38% identity) than with plant PKSs. The Cys-His-Asn catalytic triad of PKS is conserved in SbPKS with differences in some of the residues lining the active and CoA binding sites. The wild-type SbPKS displayed broad starter substrate specificity to aliphatic long-chain acyl-CoAs (C6–C14) to produce tri- and tetraketide pyrones. Mutations at H331 and N364 caused complete loss of its activity, thus suggesting that these two residues are the catalytic residues for SbPKS as in other type III PKSs. Furthermore, H227G, H227G/L366V substitutions resulted in increased tetraketide-forming activity, while wild-type SbPKS produces triketide α-pyrone as a major product. On the other hand, mutant H227G/L366V/F93A/V95A demonstrated a dramatic decrease of tetraketide pyrone formation. These observations suggest that His227 and Leu366 play an important role for the polyketide elongation reaction in SbPKS. The conformational changes in protein structure especially the cavity of the active site may have more significant effect to the activity of SbPKS compared with changes in individual residues.
Starting Page	845
Ending Page	856
Page Count	12
File Format	PDF
ISSN	14362228
Journal	Marine Biotechnology
Volume Number	13
Issue Number	5
e-ISSN	14362236
Language	English
Publisher	Springer-Verlag
Publisher Date	2010-12-23
Publisher Place	New York
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Brown seaweed Kinetics Sargassum binderi Site mutagenesis Type III polyketide synthase Freshwater & Marine Ecology Microbiology Engineering Zoology
Content Type	Text
Resource Type	Article
Subject	Aquatic Science Biotechnology

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