title: Initiation of rubber biosynthesis: In vitro comparisons of benzophenone-modified diphosphate analogues in three rubber-producing species.
abstract: Natural rubber, cis-1,4-polyisoprene, is a vital industrial material synthesized by plants via a side branch of the isoprenoid pathway by the enzyme rubber transferase. While the specific structure of this enzyme is not yet defined, based on activity it is probably a cis-prenyl transferase. Photoactive functionalized substrate analogues have been successfully used to identify isoprenoid-utilizing enzymes such as cis- and trans-prenyltransferases, and initiator binding of an allylic pyrophosphate molecule in rubber transferase has similar features to these systems. In this paper, a series of benzophenone-modified initiator analogues were shown to successfully initiate rubber biosynthesis in vitro in enzymatically-active washed rubber particles from Ficus elastica, Heveabrasiliensis and Parthenium argentatum. Rubber transferases from all three species initiated rubber biosynthesis most efficiently with farnesyl pyrophosphate. However, rubber transferase had a higher affinity for benzophenone geranyl pyrophosphate (Bz-GPP) and dimethylallyl pyrophosphate (Bz-DMAPP) analogues with ether-linkages than the corresponding GPP or DMAPP. In contrast, ester-linked Bz-DMAPP analogues were less efficient initiators than DMAPP. Thus, rubber biosynthesis depends on both the size and the structure of Bz-initiator molecules. Kinetic studies thereby inform selection of specific probes for covalent photolabeling of the initiator binding site of rubber transferase.
authors.affiliation: University of Nevada, Department of Biochemistry and Molecular Biology/MS200, Reno, NV 89557, United States.
authors.forename: Wenshuang
authors.initials: W
authors.lastname: Xie
authors.forename: Colleen M
authors.initials: CM
authors.lastname: McMahan
authors.forename: Amanda J
authors.initials: AJ
authors.lastname: Degraw
authors.forename: Mark D
authors.initials: MD
authors.lastname: Distefano
authors.forename: Katrina
authors.initials: K
authors.lastname: Cornish
authors.forename: Maureen C
authors.initials: MC
authors.lastname: Whalen
authors.forename: David K
authors.initials: DK
authors.lastname: Shintani
mesh_terms: D019659:Asteraceae / Q000378:metabolism; D001577:Benzophenones / Q000378:metabolism*; D030681:Ficus / Q000378:metabolism; D045782:Hemiterpenes / Q000096:biosynthesis* / Q000378:metabolism; D028482:Hevea / Q000378:metabolism; D007840:Latex / Q000096:biosynthesis*; D015394:Molecular Structure; D009943:Organophosphorus Compounds / Q000378:metabolism; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D012408:Rubber / Q000378:metabolism*; D012717:Sesquiterpenes / Q000378:metabolism; D013379:Substrate Specificity; D014166:Transferases / Q000378:metabolism
chemical_list: D001577:Benzophenones; D045782:Hemiterpenes; D007840:Latex; D009943:Organophosphorus Compounds; D011106:Polyisoprenyl Phosphates; D012717:Sesquiterpenes; C043060:3,3-dimethylallyl pyrophosphate; C047723:benzophenone; C015234:geranyl pyrophosphate; C004808:farnesyl pyrophosphate; C455100:1,4-polyisoprene; D012408:Rubber; D014166:Transferases; C106821:cis-prenyl transferase