Linear prenyl diphosphates whose carbon chain length varies widely from geranyl diphosphate (C
10) to natural rubber (C
>105) are biosynthesized by the catalytic function of a group of enzymes called prenyltransferases. Prenyltransferases are classified in two major groups,
trans- or (
E)-prenyltransferases and
cis- or (
Z)-prenyltransferases. From the year of 1987, many genes encoding (
E)-prenyltransferases have been cloned and characterized well. However, the structure and detailed mechanism of (
Z)-prenyltransferase had been completely unknown until our identification of a gene encoding the undecaprenyl diphosphate synthase (UPS) from
Micrococcus luteus B-P 26. Not only the primary structure but also the tertiary structure of the UPS is quite different from those of (
E)-prenyltransferases. Multiple alignment of primary structures of (
Z)-prenyltransferases identified from various organisms reveals that there are five highly conserved regions among (
Z)-prenyltransferases. In order to elucidate the biosynthetic pathway of natural rubber which is composed of
cis-1, 4-polyisoprene, we isolated and characterized two genes encoding (
Z)-prenyltransferases from the latex in
Hevea brasiliensis by using sequence information on the conserved regions of (
Z)-prenyltransferases.
In vitro rubber transferase assay using the recombinant gene product revealed that the enzyme catalyzed the formation of polyprenyl products with approximate sizes around 1×10
4 Da. Moreover, in the presence of washed bottom fraction from latex, the rubber transferase activity and the size of the major product was increased, suggesting a possible requirement of certain activation factors in the washed bottom fraction for the production of high molecular weight rubber.
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