This study investigated the enzymatic function of two putative plant GPXs, GPXle1 from Lycopersicon esculentum and GPXha2 from Helianthus annuus, which show sequence identities with the mammalian phospholipid hydroperoxide glutathione peroxidase (PHGPX). Both purified recombinant proteins expressed in Escherichia coli show PHGPX activity by reducing alkyl, fatty acid and phospholipid hydroperoxides but not hydrogen peroxide in the presence of glutathione. Interestingly, both recombinant GPXle1 and GPXha2 proteins also reduce alkyl, fatty acid and phospholipid hydroperoxides as well as hydrogen peroxide using thioredoxin as reducing substrate. Moreover, thioredoxin peroxidase (TPX) activities were found to be higher than PHGPX activities in terms of efficiency and substrate affinities, as revealed by their respective V max and K m values. We therefore conclude that these two plant GPX-like proteins are antioxidant enzymes showing PHGPX and TPX activities.

Interactions of a collection of monoclonal antibodies (mAbs) to the recombinant Nicotiana tabacum auxinbinding protein 1 (Nt-abp1) were extensively characterized using surface plasmon resonance. Dynamic interaction studies using combinations of Nt-abp1, synthetic peptides corresponding to conserved sequences within auxin-binding proteins, and the mAbs have shown that a number of the mAbs recognized discontinuous epitopes revealing the junction of distinct domains in the folded protein. In particular, the two putative auxin binding domains and the C terminus of the protein were shown to interact with each other in the folded protein. Using the auxin-induced electrical response of tobacco protoplasts as a functional assay, all the mAbs exhibited either auxin antagonist or hormonomimetic properties. These effects, measured for the first time in homologous conditions, confirm that Nt-abp1 is present at the plasma membrane and is involved in the activation of the auxin-dependent electrical response of tobacco protoplasts. Based on our surface plasmon resonance data, we propose that the key event leading to the activation of this auxin electrical response consists of a conformational change in Nt-abp1.

The dynamic mechanical properties of sisal fiber reinforced polyester composites fabricated by resin transfer molding (RTM) were investigated as a function of fiber content, frequency, and temperature. Investigation proved that at all temperature range the storage modulus (E′) value is maximum for the composites having fiber loading of 40 vol%. The loss modulus (E″) and damping peaks (tan δ) were lowered with increasing fiber content. The height of the damping peaks depends upon the fiber content and the fiber/matrix adhesion. The extent of the reinforcement was estimated from the experimental storage modulus, and it has been found that the effect of reinforcement is maximum at 40 vol% fiber content. As the fiber content increases the Tg from tan δ curve showed a positive shift. The loss modulus, storage modulus, and damping peaks were evaluated as a function of frequency. The activation energy for the glass transition increases upon the fiber content. Cole–Cole analysis was made to understand the phase behavior of the fiber reinforced composites. Finally, attempts were made to correlate the experimental dynamic properties with theoretical predictions. POLYM. COMPOS., 2009. © 2008 Society of Plastics Engineers

Phenotypical alterations observed in rolB-transformed plants have been proposed to result from a rise in intracellular free auxin due to a RolB-catalyzed hydrolysis of auxin conjugates (J.J. Estruch, J. Schell, A. Spena [1991] EMBO J 10: 3125-3128). We have investigated this hypothesis in detail using tobacco (Nicofiana tabacum) mesophyll protoplasts isolated from plants transformed with the rolB gene under the control of its own promoter (BBCUS 6 clone) or the cauliflower mosaic virus 35s promoter (CaMVBT 3 clone). Protoplasts expressing rolB showed an increased sensitivity to the auxin-induced hyperpolarization of the plasma membrane when triggered with exogenous auxin. Because this phenotypical trait was homogeneously displayed over the entire population, protoplasts were judged to be a more reliable test system than the tissue fragments used in previous studies to monitor rolB gene effects on cellular auxin levels. Accumulation of free 1-[3H]-naphthaleneacetic acid (NAA) was equivalent in CaMVBT 3, BBCUS 6, and wild-type protoplasts. Naphthyl-B-glucose ester, the major NAA metabolite in protoplasts, reached similar levels in CaMVBT 3 and normal protoplasts and was hydrolyzed at the same rate in BBCUS 6 and normal protoplasts. Furthermore, NAA accumulation and metabolism in BBCUS 6 protoplasts were independent of the rolB gene expression level. Essentially similar results were obtained with indoleacetic acid. Thus, i t was concluded that the rolB-dependent behavior of transgenic tobacco protoplasts is not a consequence of modifying the intracellular auxin concentration but likely results from changes in the auxin perception pathway.

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Hyperpolarization of tobacco protoplasts is amongst the earliest auxin responses described. It has been proposed that the auxin-binding protein, ABP1, or a related protein could be involved in the first step of auxin perception at the plasma membrane. Using for the first time homologous conditions for interaction between the protein Nt-ERabp1 or a synthetic peptide corresponding to the C-terminus and tobacco protoplasts, we have demonstrated that both can induce the hyperpolarization response. The results show that Nt-ERabp1 or the C-terminal peptide alone activates the auxin pathway from the outer face of the plasma membrane.z 1999 Federation of European Biochemical Societies.

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