Identification and reconstitution of the rubber biosynthetic machinery on rubber particles from Hevea brasiliensis

Natural rubber (NR) is stored in latex as rubber particles (RPs), rubber molecules surrounded by a lipid monolayer. Rubber transferase (RTase), the enzyme responsible for NR biosynthesis, is believed to be a member of the -prenyltransferase (cPT) family. However, none of the recombinant cPTs have sh...

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Published in:eLife 2016-10, Vol.5
Main Authors: Yamashita, Satoshi, Yamaguchi, Haruhiko, Waki, Toshiyuki, Aoki, Yuichi, Mizuno, Makie, Yanbe, Fumihiro, Ishii, Tomoki, Funaki, Ayuta, Tozawa, Yuzuru, Miyagi-Inoue, Yukino, Fushihara, Kazuhisa, Nakayama, Toru, Takahashi, Seiji
Format: Article
Language:English
Subjects:
Alcohol
Biochemistry
Biosynthesis
Biosynthetic Pathways
Euphorbiaceae
Genomics
Hevea
Hevea - genetics
Hevea - metabolism
Hevea brasiliensis
Latex
natural rubber
Observations
Peptides
Physical properties
Physiological aspects
Plant Biology
Plant Proteins - analysis
Plant Proteins - genetics
Polyclonal antibodies
Polymers
prenyltransferase
Proteins
Proteome - analysis
Proteomics
Rubber
Rubber - metabolism
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Summary:Natural rubber (NR) is stored in latex as rubber particles (RPs), rubber molecules surrounded by a lipid monolayer. Rubber transferase (RTase), the enzyme responsible for NR biosynthesis, is believed to be a member of the -prenyltransferase (cPT) family. However, none of the recombinant cPTs have shown RTase activity independently. We show that HRT1, a cPT from , exhibits distinct RTase activity in vitro only when it is introduced on detergent-washed RPs (WRPs) by a cell-free translation-coupled system. Using this system, a heterologous cPT from also exhibited RTase activity, indicating proper introduction of cPT on RP is the key to reconstitute active RTase. RP proteomics and interaction network analyses revealed the formation of the protein complex consisting of HRT1, rubber elongation factor (REF) and HRT1-REF BRIDGING PROTEIN. The RTase activity enhancement observed for the complex assembled on WRPs indicates the HRT1-containing complex functions as the NR biosynthetic machinery.
ISSN:2050-084X
2050-084X
DOI:10.7554/elife.19022