Part:BBa_K5327016
Flavin-containing monooxygenase FMO GS-OX1
Catalyzes the conversion of methylthioalkyl glucosinolates into methylsulfinylalkyl glucosinolates. Able to S-oxygenate both desulfo- and intact 4-methylthiobutyl glucosinolates, but no activity with methionine, dihomomethionine or 5-methylthiopentaldoxime.
Usage and Biology
Genome localization:Chromosome: 1; NC_003070.9
Expression diagram:
- Fig 1. The expression diagram of flavin-containing monooxygenase FMO GS-OX1
Corresponding enzyme structure:
- Fig 2. The corresponding enzyme structure of flavin-containing monooxygenase FMO GS-OX1
The PCR result:
- Fig 3. The PCR result of flavin-containing monooxygenase FMO GS-OX1
Design Notes
The design of the Flavin-containing monooxygenase FMO GS-OX1 gene is based on the coding sequence (CDS) from Arabidopsis thaliana and has been codon-optimized for efficient expression in Saccharomyces cerevisiae (S288C). FMO GS-OX1 encodes a flavin monooxygenase that catalyzes the conversion of methylthioalkyl glucosinolates into methylsulfinylalkyl glucosinolates.[4]This enzyme is capable of S-oxygenating both desulfo- and intact 4-methylthiobutyl glucosinolates but exhibits no activity with methionine, dihomomethionine, or 5-methylthiopentaldoxime. In this experiment, the GPD promoter (GPDpBBa_K517001) and PYK1 terminator (PYK1tBBa_K5327018) were selected to ensure high-level expression and mRNA stability in yeast. The optimized gene will be cloned into a vector and introduced into S. cerevisiae S288C through homologous recombination, followed by screening and expression validation using a knockout strain. This design aims to enhance the enzymatic activity of FMO GS-OX1 in yeast, thereby increasing the synthesis efficiency of methylsulfinylalkyl glucosinolates and optimizing yeast as a metabolic engineering platform for glucosinolate production.
Plasmid
- Fig 1. The plasmid expression of flavin-containing monooxygenase FMO GS-OX1
Source
Arabidopsis thaliana
References
- ↑ HANSEN B G, KLIEBENSTEIN D J, HALKIER B A. Identification of a flavin-monooxygenase as the S-oxygenating enzyme in aliphatic glucosinolate biosynthesis in Arabidopsis [J]. The Plant journal : for cell and molecular biology, 2007, 50(5): 902-10.
- ↑ KONG W, LI J, YU Q, et al. Two Novel Flavin-Containing Monooxygenases Involved in Biosynthesis of Aliphatic Glucosinolates [J]. Frontiers in plant science, 2016, 7: 1292.
- ↑ HANADA K, SAWADA Y, KUROMORI T, et al. Functional compensation of primary and secondary metabolites by duplicate genes in Arabidopsis thaliana [J]. Molecular biology and evolution, 2011, 28(1): 377-82.
- ↑ HANSEN B G, KLIEBENSTEIN D J, HALKIER B A. Identification of a flavin-monooxygenase as the S-oxygenating enzyme in aliphatic glucosinolate biosynthesis in Arabidopsis [J]. The Plant journal : for cell and molecular biology, 2007, 50(5): 902-10.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 453
Illegal EcoRI site found at 1124
Illegal EcoRI site found at 1223
Illegal XbaI site found at 700
Illegal PstI site found at 1034 - 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 453
Illegal EcoRI site found at 1124
Illegal EcoRI site found at 1223
Illegal NheI site found at 951
Illegal PstI site found at 1034 - 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 453
Illegal EcoRI site found at 1124
Illegal EcoRI site found at 1223
Illegal BamHI site found at 793
Illegal XhoI site found at 300
Illegal XhoI site found at 309 - 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 453
Illegal EcoRI site found at 1124
Illegal EcoRI site found at 1223
Illegal XbaI site found at 700
Illegal PstI site found at 1034 - 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 453
Illegal EcoRI site found at 1124
Illegal EcoRI site found at 1223
Illegal XbaI site found at 700
Illegal PstI site found at 1034 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 64
Illegal BsaI site found at 139
Illegal BsaI site found at 1257
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