2011 Annual Report 1a.Objectives (from AD-416) To advance the fundamental knowledge regarding structure/property relationships in natural rubber, especially the role of naturally-occurring, non-rubber constituents in creating and controlling the nanostructured features that are believed to be responsible for the unique properties of natural rubber derived from Hevea brasiliensis. 1b.Approach (from AD-416) 1) Deconstruct natural rubber latex through selective separation of naturally-occurring proteins and lipids by: a) identifying highly abundant natural rubber proteins (proteomics) b) extract and concentrate proteins from natural latex, including Hevea c) produce recombinant rubber particle proteins 2) Systematically reconstruct the latex composition from synthetic and Hevea, guayule, and Russian dandelion natural latex by protein add-back compositions. 3) Fully characterize the nanostructure and properties of resulting materials. Formerly 5325-41000-048-03S (September, 2010). 3.Progress Report In FY2011 collaborative research revealed that blends of low-protein guayule latex and high-protein Hevea latex exhibit intermediate physical properties (tensile strength, gel, and thermo-oxidative stability) related to the beneficial effects of proteins. This suggests that proteins, or their peptide and amino acid degradation products, in Hevea latex are physically and chemically capable of interacting with guayule rubber in the blend. Separately, expression and purification of Hevea REF (a 14 KDa. protein) was carried out using a pGEX-6P-1 vector. For protein expression, the REF cDNA was cloned between EcoR I and Xho I sites of pGEX-6P-1 vector to produce a recombinant plasmid with REF. Then the recombinant plasmid was transferred to E. coli competent cells (Rossetta cells). Maximum concentration of REF protein was achieved 6 hours after induction by 0.1 mM IPTG. Purification of highly-abundant Hevea proteins may elucidate their role in protein-polymer interactions. The agreement was monitored through frequent emails and telephone calls, and two face-to-face meetings in FY2011.