http://www.biochemj.org Biochemical Journal RSS feed -- BJ Plant Immediate Publications 0264-6021 1470-8728 Biochemical Journal http://www.biochemj.org/images/BJ_Name.gif http://www.biochemj.org http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111792 in vitro. A MBP kinase assay of the immunoprecipitation complex indicated that OsMPK3 and OsMPK6 were activated in response to a moderately low temperature (12°C) but a severely low temperature (4°C) in rice seedlings. A constitutively active form of OsMKK6, OsMKK6DD, showed elevated phosphorylation activity against OsMPK3 and OsMPK6 in vitro. OsMPK3, but not OsMPK6, was constitutively activated in transgenic plants overexpressing OsMKK6DD, indicating that OsMPK3 is an in vivo target of OsMKK6. Enhanced chilling tolerance was observed in the transgenic plants overexpressing OsMKK6DD. Taken together, our data suggest that OsMKK6 and OsMPK3 constitute a moderately low-temperature signaling pathway and regulate cold stress tolerance in rice.]]> G Xie, H Kato, R Imai 2012-01-16T14:27:38Z doi:10.1042/BJ20111792 Portland Press Limited 2012-01-16 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111308 AtTDP gene in Arabidopsis thaliana, an orthologue of yeast and human TDP1 genes (Lee et al (2010) Plant Physiology 154; 1460-1469). Sequence alignment of TDP1 orthologues revealed that AtTDP has both a conserved C-terminal TDP domain and, uniquely, an N-terminal SMAD/forkhead-associated (FHA) domain. To help understand the function of this novel enzyme, we analyzed the substrate saturation kinetics of full­length AtTDP versus a truncated AtTDP mutant lacking the N–terminal FHA domain. The recombinant AtTDP protein hydrolyzed a single-stranded DNA substrate with K_m and k_cat / K_m values of 703 ± 137 nM and 1.5 x 10⁹ ± 0.04 x 10⁹ M^-1 min^-1, respectively. The AtTDP D1–122 protein (TDP domain) showed kinetic parameters that were equivalent to those of the full­length AtTDP protein. A basic amino acid sequence (RKKVKP) within the AtTDP D123–605 protein (FHA domain) was necessary for nuclear localization of AtTDP. Analysis of active site mutations showed that a histidine and a lysine residue in each of the HKD motifs were critical for enzyme activity. Vanadates, inhibitors of phosphoryl transfer reactions, inhibited AtTDP enzymatic activity and retarded the growth of an Arabidopsis tdp mutant. Finally, we showed that expression of the AtTDP gene could complement a yeast tdp1Drad1D mutant, rescuing the growth inhibitory effects of vanadate analogs and camptothecin (CPT). Taken together, our data demonstrate the structure-based function of AtTDP through which AtTDP can repair DNA strand breaks in plants.]]> H Kim, S Na, S Lee, Y Jeong, H Hwang, J Hur, S Park, J Woo, S Kim 2012-01-03T16:39:38Z doi:10.1042/BJ20111308 Portland Press Limited 2012-01-03 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111796 Synechocystis sp. PCC6803 and report here on the role of the membrane-bound 4-hydroxybenzoate solanesyltransferase, Slr0926, in PQ-9 biosynthesis and on the properties of the enzyme. The catalytic activity of Slr0926 was demonstrated by in vivo labelling experiments in Synechocystis sp., complementation studies in an E. coli mutant with a defect in ubiquinone biosynthesis as well as by in vitro assays using the recombinant as well as the native enzyme. While Slr0926 was highly specific for the prenyl acceptor substrate, 4-hydroxybenzoate, it displayed a broad specificity with regard to the prenyl donor substrate and used not only solanesyl diphosphate (SPP) but also a number of shorter-chain prenyl diphosphates. In combination with in silico data, our results indicate that Slr0926 evolved from bacterial 4-hydroxybenzoate prenyltransferase catalysing prenylation in the course of ubiquinone biosynthesis.]]> R Sadre, C Pfaff, S Buchkremer 2011-12-14T11:13:05Z doi:10.1042/BJ20111796 Portland Press Limited 2011-12-14 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111739 DREB2A which lacks the interaction domain accumulates during heat stress and senescence. In addition RCD1 is rapidly degraded during heat stress, thus our results suggest that removal of RCD1 protein or the loss of the interaction domain in DREB2A appears to be required for proper DREB2A function under stress conditions.]]> J P Vainonen, P Jaspers, M Wrzaczek, A Lamminmäki, R A Reddy, L Vaahtera, M Brosché, J Kangasjärvi 2011-12-12T16:51:43Z doi:10.1042/BJ20111739 Portland Press Limited 2011-12-12 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111311 Arabidopsis. The BCD1 gene is induced by excessive iron but repressed by iron deficiency. It is also induced by cellular and tissue damages occurring under osmotic stress. The activation-tagged mutant bcd1-1D exhibits leaf chlorosis, typical symptom of iron deficiency. The chlorotic lesion of the mutant was partially recovered by iron feeding. Whereas the bcd1-1D mutant accumulated a lower amount of iron, the iron level was elevated in the knockout mutant bcd1-1. The BCD1 protein is localized to the Golgi complex. We propose that the BCD1 transporter plays a role in sustaining iron homeostasis by reallocating excess iron released from stress-induced cellular damages.]]> P Seo, J Park, M Park, Y Kim, S Kim, J Jung, C Park 2011-12-08T11:38:33Z doi:10.1042/BJ20111311 Portland Press Limited 2011-12-08 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111877 Anabaena PCC 7120 genome harbours seven genes/ORFs with homology to peroxiredoxins. One of these (all1541) was identified to encode a novel glutaredoxin (Grx) domain-containing peroxiredoxin by bioinformatic analysis. A recombinant N-terminal His-tagged All1541 protein was overexpressed in E. coli and purified. Analysis with protein alkylating agent AMS showed All1541 to form an intra-molecular disulfide bond. The All1541 protein used glutathione (GSH) more efficiently than thioredoxin (Trx) to detoxify H₂O₂. Deletion of Grx domain from All1541 resulted in loss of GSH-dependent peroxidase activity. Employing site directed mutagenesis, the cysteine residues at position 50 and 75 were identified as peroxidatic and resolving cysteine residues respectively, while both the cysteine residues within the Grx domain (position 181 and 184) were shown to be essential for GSH-dependent peroxidase activity. Based on these data, a reaction mechanism has been proposed for All1541. In vitro All1541 protein protected plasmid DNA from oxidative damage. In Anabaena PCC 7120, the all1541 was transcriptionally activated under oxidative stress. Recombinant Anabaena PCC 7120 strain over-expressing All1541 protein showed superior oxidative stress tolerance to H₂O₂ as compared to the wild-type strain. The results suggest that the glutathione dependent peroxidase All1541 plays an important role in protecting Anabaena from oxidative stress.]]> M Banerjee, A Ballal, S Kumar Apte 2011-12-08T10:28:25Z doi:10.1042/BJ20111877 Portland Press Limited 2011-12-08 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111443 in vitro measurement of Rubisco carboxylase activity prevented a progressive decline in Rubisco turnover. This was due to the removal of an inhibitory bisphosphate which was present in the RuBP preparation, and was found to be D-glycero-2,3-pentodiulose-1,5-bisphosphate (PDBP). The substrate specificity of the expressed protein indicates a role for CA1P phosphatase in the removal of misfire products of Rubisco.]]> P John Andralojc, P J Madgwick, Y Tao, A Keys, J L Ward, M H Beale, J E Loveland, P J Jackson, A C Willis, S Gutteridge, M AJ Parry 2011-12-02T11:12:27Z doi:10.1042/BJ20111443 Portland Press Limited 2011-12-02 BJ Plant http://www.biochemj.org/bj/imps/refer.htm?MSID=BJ20111378 6 M^-1 s^-1. In PtGpx5, Glu79, which replaces the Gln usually found in Gpx catalytic tetrad, is likely involved in substrate selectivity. Although the redox midpoint potential of the Cys44-Cys92 disulfide and the pKa of Cys44 are not modified in the E79Q variant, it exhibited significantly improved kinetic parameters (K_peroxide and k_cat) with tert-butyl hydroperoxide. The characterization of the monomeric Y151R variant demonstrated that PtGpx5 is not an obligate homodimer. Also, we show that the conserved Phe90 is important for Trx recognition and that Trx-mediated recycling of PtGpx5 occurs via the formation of a transient disulfide between the Trx catalytic cysteine and the Gpx5 resolving cysteine. Finally, we demonstrate that the conformational changes observed during the transition from the reduced to the oxidized form of PtGpx5 are primarily determined by the oxidation of the peroxidatic cysteine into sulfenic acid. Besides, mass spectrometry analysis of in vitro oxidized PtGpx5 demonstrated that the peroxidatic cysteine can be over-oxidized into sulfinic or sulfonic acids. This suggests that some isoforms could have dual functions potentially acting as hydrogen peroxide- and peroxynitrite-scavenging systems and/or as mediators of peroxide signalling as proposed for 2-Cys peroxiredoxins.]]> B Selles, M Hugo, M Trujillo, V Srivastava, G Wingsle, J Jacquot, R Radi, N Rouhier 2011-11-29T12:29:48Z doi:10.1042/BJ20111378 Portland Press Limited 2011-11-29 BJ Plant