Journal article
Next Generation Sequencing in Predicting Gene Function in Podophyllotoxin Biosynthesis
The Journal of biological chemistry, Vol.288(1), pp.466-479
01/04/2013
Handle:
https://hdl.handle.net/2376/116379
PMCID: PMC3537044
PMID: 23161544
Abstract
Background:
Biosynthetic pathways to structurally complex plant medicinals are incomplete or unknown.
Results:
Next generation sequencing/bioinformatics and metabolomics analysis of
Podophyllum
tissues gave putative unknown genes in podophyllotoxin biosynthesis.
Conclusion:
Regio-specific methylenedioxy bridge-forming CyP450s were identified catalyzing pluviatolide formation.
Significance:
Database of several medicinal plant transcriptome assemblies and metabolic profiling are made available for scientific community.
Podophyllum
species are sources of (−)-podophyllotoxin, an aryltetralin lignan used for semi-synthesis of various powerful and extensively employed cancer-treating drugs. Its biosynthetic pathway, however, remains largely unknown, with the last unequivocally demonstrated intermediate being (−)-matairesinol. Herein, massively parallel sequencing of
Podophyllum hexandrum
and
Podophyllum peltatum
transcriptomes and subsequent bioinformatics analyses of the corresponding assemblies were carried out. Validation of the assembly process was first achieved through confirmation of assembled sequences with those of various genes previously established as involved in podophyllotoxin biosynthesis as well as other candidate biosynthetic pathway genes. This contribution describes characterization of two of the latter, namely the cytochrome P450s, CYP719A23 from
P. hexandrum
and CYP719A24 from
P. peltatum
. Both enzymes were capable of converting (−)-matairesinol into (−)-pluviatolide by catalyzing methylenedioxy bridge formation and did not act on other possible substrates tested. Interestingly, the enzymes described herein were highly similar to methylenedioxy bridge-forming enzymes from alkaloid biosynthesis, whereas candidates more similar to lignan biosynthetic enzymes were catalytically inactive with the substrates employed. This overall strategy has thus enabled facile further identification of enzymes putatively involved in (−)-podophyllotoxin biosynthesis and underscores the deductive power of next generation sequencing and bioinformatics to probe and deduce medicinal plant biosynthetic pathways.
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Details
- Title
- Next Generation Sequencing in Predicting Gene Function in Podophyllotoxin Biosynthesis
- Creators
- Joaquim V Marques - From theKye-Won Kim - From theChoonseok Lee - From theMichael A Costa - From theGregory D May - theJohn A Crow - theLaurence B Davin - From theNorman G Lewis - From the
- Publication Details
- The Journal of biological chemistry, Vol.288(1), pp.466-479
- Academic Unit
- Biological Chemistry, Institute of
- Publisher
- American Society for Biochemistry and Molecular Biology; 9650 Rockville Pike, Bethesda, MD 20814, U.S.A
- Grant note
- 1RC2GM092561 / National Institutes of Health
- Identifiers
- 99900548211301842
- Language
- English
- Resource Type
- Journal article