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Ziegler, J.; Abel S. Analysis of amino acids by HPLC/electrospray negative ion tandem mass spectrometry using 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl) derivatization Amino Acids 46, 2799-2808, (2014) DOI: 10.1007/s00726-014-1837-5

A new method for the determination of amino acids is presented. It combines established methods for the derivatization of primary and secondary amino groups with 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl) with the subsequent amino acid specific detection of the derivatives by LC–ESI–MS/MS using multiple reaction monitoring (MRM). The derivatization proceeds within 5 min, and the resulting amino acid derivatives can be rapidly purified from matrix by solid-phase extraction (SPE) on HR-X resin and separated by reversed-phase HPLC. The Fmoc derivatives yield several amino acid specific fragment ions which opened the possibility to select amino acid specific MRM transitions. The method was applied to all 20 proteinogenic amino acids, and the quantification was performedusing l-norvaline as standard. A limit of detection as low as 1 fmol/μl with a linear range of up to 125 pmol/μl could be obtained. Intraday and interday precisions were lower than10 % relative standard deviations for most of the amino acids. Quantification usingl-norvaline as internal standard gave very similar results compared to the quantificationusing deuterated amino acid as internal standards. Using this protocol, it was possible to record the amino acid profiles of only a single root from Arabidopsis thaliana seedlings and to compare it with the amino acid profiles of 20 dissected root meristems (200 μm).

Calderón Villalobos, L. I. A.; Lee, S.; De Oliveira, C.; Ivetac, A.; Brandt, W.; Armitage, L.; Sheard, L. B.; Tan, X.; Parry, G.; Mao, H.; Zheng, N.; Napier, R.; Kepinski, S.; Estelle, M. A combinatorial TIR1/AFB–Aux/IAA co-receptor system for differential sensing of auxin Nat Chem Biol 8, 477-485, (2012) DOI: 10.1038/nchembio.926

The plant hormone auxin regulates virtually every aspect of plant growth and development. Auxin acts by binding the F-box protein transport inhibitor response 1 (TIR1) and promotes the degradation of the AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) transcriptional repressors. Here we show that efficient auxin binding requires assembly of an auxin co-receptor complex consisting of TIR1 and an Aux/IAA protein. Heterologous experiments in yeast and quantitative IAA binding assays using purified proteins showed that different combinations of TIR1 and Aux/IAA proteins form co-receptor complexes with a wide range of auxin-binding affinities. Auxin affinity seems to be largely determined by the Aux/IAA. As there are 6 TIR1/AUXIN SIGNALING F-BOX proteins (AFBs) and 29 Aux/IAA proteins in Arabidopsis thaliana, combinatorial interactions may result in many co-receptors with distinct auxin-sensing properties. We also demonstrate that the AFB5Aux/IAA co-receptor selectively binds the auxinic herbicide picloram. This co-receptor system broadens the effective concentration range of the hormone and may contribute to the complexity of auxin response.

Fonseca, S.; Chini, A.; Hamberg, M.; Adie, B.; Porzel, A.; Kramell, R.; Miersch, O.; Wasternack, C.; Solano, R. (+)-7-iso-Jasmonoyl-L-isoleucine is the endogenous bioactive jasmonate Nat Chem Biol 5, 344-350, (2009) DOI: 10.1038/nchembio.161

Hormone-triggered activation of the jasmonate signaling pathway in Arabidopsis thaliana requires SCFCOI1-mediated proteasome degradation of JAZ repressors. (-)-JA-L-Ile is the proposed bioactive hormone, and SCFCOI1 is its likely receptor. We found that the biological activity of (-)-JA-L-Ile is unexpectedly low compared to coronatine and the synthetic isomer (+)-JA-L-Ile, which suggests that the stereochemical orientation of the cyclopentanone-ring side chains greatly affects receptor binding. Detailed GC-MS and HPLC analyses showed that the (-)-JA-L-Ile preparations currently used in ligand binding studies contain small amounts of the C7 epimer (+)-7-iso-JA-L-Ile. Purification of each of these molecules demonstrated that pure (-)-JA-L-Ile is inactive and that the active hormone is (+)-7-iso-JA-L-Ile, which is also structurally more similar to coronatine. In addition, we show that pH changes promote conversion of (+)-7-iso-JA-L-Ile to the inactive (-)-JA-L-Ile form, thus providing a simple mechanism that can regulate hormone activity through epimerization.

Santner, A.; Calderón Villalobos, L.I.; Estelle, M. Plant hormones are versatile chemical regulators of plant growth Nat Chem Biol 5(5), 301-307, (2009) DOI: 10.1038/nchembio.165

The plant hormones are a structurally unrelated collection of small molecules derived from various essential metabolic pathways. These compounds are important regulators of plant growth and mediate responses to both biotic and abiotic stresses. During the last ten years there have been many exciting advances in our understanding of plant hormone biology, including new discoveries in the areas of hormone biosynthesis, transport, perception and response. Receptors for many of the major hormones have now been identified, providing new opportunities to study the chemical specificity of hormone signaling. These studies also reveal a surprisingly important role for the ubiquitin-proteasome pathway in hormone signaling. In addition, recent work confirms that hormone signaling interacts at multiple levels during plant growth and development. In the future, a major challenge will be to understand how the information conveyed by these simple compounds is integrated during plant growth.
Bücher und Buchkapitel

Vaira, A.M.; Acotto, G.P.; Gago-Zachert, S.; García, M.L.; Grau, O.; Milne, R.G.; Morikawa, T.; Natsuaki, T.; Torov, V.; Verbeek, M.; Vetten, H.J. Genus Ophiovirus (Fauquet, C. M., Mayo, M. A., Maniloff, J., Desselberger, U., Ball, L. A.). Elsevier, Academic Press 673-679, (2005) ISBN: 9780080575483; 9780122499517

Virus Taxonomy is a standard and comprehensive source for the classification of viruses, created by the International Committee of the Taxonomy of Viruses. The book includes eight taxonomic reports of the ICTV and provides comprehensive information on 3 taxonomic orders of viruses, 73 families, 9 subfamilies, 287 genera, and 1938 virus species. The book also features about 429 colored pictures and diagrams for more efficient learning. The text is divided into four parts, comprised of 16 chapters and presenting the following features: • Compiled data from numerous international experts about virus taxonomy and nomenclature • Organized information on over 6000 recognized viruses, illustrated with diagrams of genome organization and virus replication cycle • Data on the phylogenetic relationships among viruses of the same and different taxa • Discussion of the qualitative and quantitative relationships of virus sequences The book is a definitive reference for microbiologists, molecular biologists, research-level virologists, infectious disease specialists, and pharmaceutical researchers working on antiviral agents. Students and novices in taxonomy and nomenclature will also find this text useful. 

Abel, S.; Savchenko, T.; Levy, M. Genome-wide comparative analysis of the <em>IQD</em> gene families in <em>Arabidopsis thaliana</em> and Oryza sativa BMC Evolutionary Biology 5, 72 (1-25), (2005)

We identified and analyzed 33 and 29 IQD1-like genes in Arabidopsis thaliana and Oryza sativa, respectively. The encoded IQD proteins contain a plant-specific domain of 67 conserved amino acid residues, referred to as the IQ67 domain, which is characterized by a unique and repetitive arrangement of three different calmodulin recruitment motifs, known as the IQ, 1-5-10, and 1-8-14 motifs. We demonstrated calmodulin binding for IQD20, the smallest IQD protein in Arabidopsis, which consists of a C-terminal IQ67 domain and a short N-terminal extension. A striking feature of IQD proteins is the high isoelectric point (~10.3) and frequency of serine residues (~11%). We compared the Arabidopsis and rice IQD gene families in terms of gene structure, chromosome location, predicted protein properties and motifs, phylogenetic relationships, and evolutionary history. The existence of an IQD-like gene in bryophytes suggests that IQD proteins are an ancient family of calmodulin-binding proteins and arose during the early evolution of land plants. Comparative phylogenetic analyses indicate that the major IQD gene lineages originated before the monocot-eudicot divergence. The extant IQD loci in Arabidopsis primarily resulted from segmental duplication and reflect preferential retention of paralogous genes, which is characteristic for proteins with regulatory functions. Interaction of IQD1 and IQD20 with calmodulin and the presence of predicted calmodulin binding sites in all IQD family members suggest that IQD proteins are a new class of calmodulin targets. The basic isoelectric point of IQD proteins and their frequently predicted nuclear localization suggest that IQD proteins link calcium signaling pathways to the regulation of gene expression. Our comparative genomics analysis of IQD genes and encoded proteins in two model plant species provides the first step towards the functional dissection of this emerging family of putative calmodulin targets.
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