Rubber Molecular Weight Regulation, in Vitro, in Plant Species that Produce High and Low Molecular Weights in Vivo

Abstract

In three rubber-producing species, in vitro, the rates of initiation and polymerization and the biopolymer molecular weight produced were affected by the concentration of farnesyl diphosphate (FPP) initiator and isopentenyl diphosphate (IPP) elongation substrate (monomer). Ficus elastica, a low molecular weight-producer in vivo, synthesized rubber polymers approximately twice the molecular weight of those made by Hevea brasiliensis or Parthenium argentatum (which produce high molecular weights in vivo), possibly due to its lower IPP Km. In all species, increasing FPP concentrations increased rubber biosynthetic rate and new molecules initiated but decreased molecular weight by competition with the allylic diphosphate (APP) end of elongating rubber molecules for the APP binding site. Increasing IPP concentrations increased rubber biosynthetic rate and rubber molecular weight, but only when FPP concentrations were below the FPP Km's or where negative cooperativity operated. In conclusion, rubber transferase is not the prime regulator of rubber molecular weight in vivo.

Natural rubber (NR) biosynthesis: perspectives from polymer chemistry

“…importantly, they demonstrated that increasing the amount of iPP while keeping the FPP concentration constant resulted in increased MW in three different types of WrPs: Hevea, guayule and Ficus elastica (Fig. 2.17) (Castillón and Cornish, 1999;Cornish et al, 2000). These findings support the notion of a living-like polymerization.…”

Molecular Cloning and Characterization of Rubber Biosynthetic Genes from Taraxacum koksaghyz

“…Natural rubber is composed of cis-1,4 polyisoprene with molecular weights ranging from <50 kDa in Ficus elastica to >1,000 kDa in H. brasiliensis (Cornish 1993;Castillón and Cornish 1999;Cornish et al 2000). Rubber biosynthesis takes place on the surface of rubber particles and is catalyzed by rubber-particle-bound cis-prenyltranferases (CPTs) via cis-1,4-polymerization of isoprene monomers derived from isopentenyl diphosphate (IPP).…”

Protein farnesyltransferase inhibitors interfere with farnesyl diphosphate binding by rubber transferase

“…On the other hand, the noncompetitive inhibition by 6 of H. brasiliensis rubber transferase may be caused by interference resulting from the additional binding of compound 6 at the IPP binding site, which is in the proximity of the allylic diphosphate binding site. Competition between IPP and allylic diphosphate for the IPP binding sites has been observed at high substrate concentrations [1,16], and 6 was present at a range of 2500 to 10 000‐fold molar excess to FPP in the assay (30‐ to 130‐times the K m for FPP [17]). These results suggest that the spatial orientation between the IPP and initiator binding sites differs between the two rubber transferases, as well as the ability to bind 6 at other surfaces within the active site.…”

“…The different behavior between the two rubber transferases towards 6 should also be considered in light of the cooperative effects between IPP and FPP if one assumes that 6 is perceived as FPP by both enzymes. In the range of concentrations of 6 incubated in the assays, the two rubber transferases exhibit different degrees of negative cooperativity, with the P. argentatum enzyme showing the strongest effect [17]. Under these conditions, binding of the first initiator molecule decreases the ability of a free FPP to displace the bound FPP (or elongating polymer).…”

“…The ability of the FPP‐like compound 6 to bind noncompetitively at an additional location within the H. brasiliensis active site may explain some of differences in the degree of negative cooperativity between the two enzymes. Furthermore, the development of the P. argentatum rubber transferase with a low K m for FPP (about 150‐fold lower in concentration than the corresponding constant for H. brasiliensis [17]) may also explain the difference in degree of negative cooperativity, because compound 6 can bind competitively into the initiator site of the P. argentatum enzyme, unlike the noncompetitive binding in the vicinity of the weaker affinity initiator binding site of the H. brasiliensis protein. More experimentation to elucidate the mechanism of the negative cooperativity can be performed, but the basis for this behavior will probably only become apparent after the crystal structures of the rubber transferases have been determined.…”