title: Identification and reconstitution of the rubber biosynthetic machinery on rubber particles fromHevea brasiliensis.
abstract: 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 thecis-prenyltransferase (cPT) family. However, none of the recombinant cPTs have shown RTase activity independently. We show that HRT1, a cPT fromHeveabrasiliensis, exhibits distinct RTase activity in vitro only when it is introduced on detergent-washedHeveaRPs (WRPs) by a cell-free translation-coupled system. Using this system, a heterologous cPT fromLactucasativaalso 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.
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Satoshi
authors: S
authors: Yamashita
authors: Sumitomo Rubber Industries, Ltd, Kobe, Japan.
authors: Haruhiko
authors: H
authors: Yamaguchi
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Toshiyuki
authors: T
authors: Waki
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Yuichi
authors: Y
authors: Aoki
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Makie
authors: M
authors: Mizuno
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Fumihiro
authors: F
authors: Yanbe
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Tomoki
authors: T
authors: Ishii
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Ayuta
authors: A
authors: Funaki
authors: Graduate School of Science and Engineering, Saitama University, Saitama, Japan.
authors: Yuzuru
authors: Y
authors: Tozawa
authors: Sumitomo Rubber Industries, Ltd, Kobe, Japan.
authors: Yukino
authors: Y
authors: Miyagi-Inoue
authors: Sumitomo Rubber Industries, Ltd, Kobe, Japan.
authors: Kazuhisa
authors: K
authors: Fushihara
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Toru
authors: T
authors: Nakayama
authors: Graduate School of Engineering, Tohoku University, Sendai, Japan.
authors: Seiji
authors: 0000-0002-2288-4340
authors: S
authors: Takahashi
mesh_terms: D053898:Biosynthetic Pathways; D028482:Hevea / Q000235:genetics* / Q000378:metabolism*; D010940:Plant Proteins / Q000032:analysis / Q000235:genetics; D020543:Proteome / Q000032:analysis; D012408:Rubber / Q000378:metabolism*
chemical_list: D010940:Plant Proteins; D020543:Proteome; D012408:Rubber
keywords: E. coli; Hevea brasiliensis ; S. cerevisiae; biochemistry; natural rubber; plant biology; prenyltransferase