Enzymatic Properties and Mutational Studies of Chalcone Synthase from Physcomitrella patens

Raja Noor Zaliha Raja Abdul Rahman,Mahiran Basri,Iffah Izzati Zakaria,Abu Bakar Salleh

doi:10.3390/ijms13089673

Raja Noor Zaliha Raja Abdul Rahman, Mahiran Basri + Show 2 more

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https://doi.org/10.3390/ijms13089673

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Abstract

PpCHS is a member of the type III polyketide synthase family and catalyses the synthesis of the flavonoid precursor naringenin chalcone from p-coumaroyl-CoA. Recent research reports the production of pyrone derivatives using either hexanoyl-CoA or butyryl-CoA as starter molecule. The Cys-His-Asn catalytic triad found in other plant chalcone synthase predicted polypeptides is conserved in PpCHS. Site directed mutagenesis involving these amino acids residing in the active-site cavity revealed that the cavity volume of the active-site plays a significant role in the selection of starter molecules as well as product formation. Substitutions of Cys 170 with Arg and Ser amino acids decreased the ability of the PpCHS to utilize hexanoyl-CoA as a starter molecule, which directly effected the production of pyrone derivatives (products). These substitutions are believed to have a restricted number of elongations of the growing polypeptide chain due to the smaller cavity volume of the mutant’s active site.

Highlights

Flavonoids and isoflavonoids are important plant secondary metabolites that mediate diverse biological functions such as chemical messengers and antioxidants
The recombinant Physcomitrella patens chalcone synthase was purified to homogeneity through a single step of affinity chromatography using Ni2+ Sepharose and characterized
The work presented here has highlighted the profile of chalcone synthase enzymatic properties based on the production of a product using hexanoyl-CoA as a substrate

Summary

Introduction

Flavonoids and isoflavonoids are important plant secondary metabolites that mediate diverse biological functions such as chemical messengers and antioxidants. One of the main key enzymes involved in the flavonoid biosynthesis is chalcone synthase (CHS). The synthesis of naringenin chalcone, from which diverse flavonoid end products are derived, led to extensive studies that have been done involving hundreds of CHS genes cloned and characterized from different types of plants [2,3]. Mosses (Musci, Bryophytae) are one of the oldest groups of land plants. Their life cycle mainly consists of a simple photoautotrophic haploid gametophytic generation. The potential of mosses as a model system in studying higher plant biological processes has led to the first genomic sequence of Physcomitrella patens [4]. In this study, the CHS gene (Acession No: DQ 275627.2) was cloned from the gametophore tissues of P. patens due to the availability of the P. patens EST and genomic sequences

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