Impact of methionine nutrition on the leaf proteome of Lupinus angustifolius L. and Vicia faba L.

Grain legumes possess a gernerally favourable seed protein amino acid composition but low contents of sulfur amino acids decrease their nutrient value. To enhance the methionie content of local grain legumes by plant breeding, phenotypical attributes for high methionine contents are required for the selection process

Structure and function of the mitochondrial NADH ubiquinone oxidoreductase of Arabidopsis thaliana

[no abstract

Defining the protein complex proteome of plant mitochondria

A classical approach, protein separation by two-dimensional blue native/sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was combined with tandem mass spectrometry and up-to-date computer technology to characterize the mitochondrial "protein complex proteome" of Arabidopsis (Arabidopsis thaliana) in so far unrivaled depth. We further developed the novel GelMap software package to annotate and evaluate two-dimensional blue native/sodium dodecyl sulfate gels. The software allows (1) annotation of proteins according to functional and structural correlations (e.g. subunits of a distinct protein complex), (2) assignment of comprehensive protein identification lists to individual gel spots, and thereby (3) selective display of protein complexes of low abundance. In total, 471 distinct proteins were identified by mass spectrometry, several of which form part of at least 35 different mitochondrial protein complexes. To our knowledge, numerous protein complexes were described for the first time (e.g. complexes including pentatricopeptide repeat proteins involved in nucleic acid metabolism). Discovery of further protein complexes within our data set is open to everybody via the public GelMap portal at www.gelmap.de/arabidopsis_mito

Supramolecular structure of the OXPHOS system in highly thermogenic tissue of Arum maculatum

The protein complexes of the mitochondrial respiratory chain associate in defined ways forming supramolecular structures called respiratory supercomplexes or respirasomes. In plants, additional oxidoreductases participate in respiratory electron transport, e.g. the so-called "alternative NAD(P)H dehydrogenases" or an extra terminal oxidase called "alternative oxidase" (AOX). These additional enzymes were previously reported not to form part of respiratory supercomplexes. However, formation of respiratory supercomplexes might indirectly affect "alternative respiration" because electrons can be channeled within the supercomplexes which reduces access of the alternative enzymes towards their electron donating substrates. Here we report an investigation on the supramolecular organization of the respiratory chain in thermogenic Arum maculatum appendix mitochondria, which are known to have a highly active AOX for heat production. Investigations based on mild membrane solubilization by digitonin and protein separation by blue native PAGE revealed a very special organization of the respiratory chain in A. maculatum, which strikingly differs to the one described for the model plant Arabidopsis thaliana: (i) complex I is not present in monomeric form but exclusively forms part of a I + III2 supercomplex, (ii) the III2 + IV and I + III2 + IV supercomplexes are detectable but of low abundance, (iii) complex II has fewer subunits than in A. thaliana, and (iv) complex IV is mainly present as a monomer in a larger form termed "complex IVa". Since thermogenic tissue of A. maculatum at the same time has high AOX and I + III2 supercomplex abundance and activity, negative regulation of the alternative oxidase by supercomplex formation seems not to occur. Functional implications are discussed. © 2010 Elsevier Masson SAS. All rights reserved

Involvement of the Azotobacter vinelandii Rhodanese-Like Protein RhdA in the Glutathione Regeneration Pathway

The phenotypic features of the Azotobacter vinelandii RhdA mutant MV474 (in which the rhdA gene was deleted) indicated that defects in antioxidant systems in this organism were related to the expression of the tandem-domain rhodanese RhdA. In this work, further insights on the effects of the oxidative imbalance generated by the absence of RhdA (e.g. increased levels of lipid hydroperoxides) are provided. Starting from the evidence that glutathione was depleted in MV474, and using both in silico and in vitro approaches, here we studied the interaction of wild-type RhdA and Cys(230) Ala site-directed RhdA mutant with glutathione species. We found that RhdA was able to bind in vitro reduced glutathione (GSH) and that RhdA-Cys(230) residue was mandatory for the complex formation. RhdA catalyzed glutathione-disulfide formation in the presence of a system generating the glutathione thiyl radical (GS(.), an oxidized form of GSH), thereby facilitating GSH regeneration. This reaction was negligible when the Cys230 Ala RhdA mutant was used. The efficiency of RhdA as catalyst in GS(.)-scavenging activity is discussed on the basis of the measured parameters of both interaction with glutathione species and kinetic studies.Vigoni project/0815171Deutscher Akademischer Austausch DienstUniversita` degli Studi di Milano/Fondo interno ricerca scientifica e tecnologic

L-galactono-1,4-lactone dehydrogenase (GLDH) forms part of three subcomplexes of mitochondrial complex I in Arabidopsis thaliana

L-Galactono-1,4-lactone dehydrogenase (GLDH) catalyzes the terminal step of the Smirnoff-Wheeler pathway for vitamin C (L-ascorbate) biosynthesis in plants. A GLDH in gel activity assay was developed to biochemically investigate GLDH localization in plant mitochondria. It previously has been shown that GLDH forms part of an 850-kDa complex that represents a minor form of the respiratory NADH dehydrogenase complex (complex I). Because accumulation of complex I is disturbed in the absence of GLDH, a role of this enzyme in complex I assembly has been proposed. Here we report that GLDH is associated with two further protein complexes. Using native gel electrophoresis procedures in combination with the in gel GLDH activity assay and immunoblotting, two mitochondrial complexes of 470 and 420 kDa were identified. Both complexes are of very low abundance. Protein identifications by mass spectrometry revealed that they include subunits of complex I. Finally, the 850-kDa complex was further investigated and shown to include the complete "peripheral arm" of complex I. GLDH is attached to a membrane domain, which represents a major fragment of the "membrane arm" of complex I. Taken together, our data further support a role of GLDH during complex I formation, which is based on its binding to specific assembly intermediates.Instituto de Fisiología Vegeta

L-galactono-1,4-lactone dehydrogenase (GLDH) forms part of three subcomplexes of mitochondrial complex I in Arabidopsis thaliana

L-Galactono-1,4-lactone dehydrogenase (GLDH) catalyzes the terminal step of the Smirnoff-Wheeler pathway for vitamin C (L-ascorbate) biosynthesis in plants. A GLDH in gel activity assay was developed to biochemically investigate GLDH localization in plant mitochondria. It previously has been shown that GLDH forms part of an 850-kDa complex that represents a minor form of the respiratory NADH dehydrogenase complex (complex I). Because accumulation of complex I is disturbed in the absence of GLDH, a role of this enzyme in complex I assembly has been proposed. Here we report that GLDH is associated with two further protein complexes. Using native gel electrophoresis procedures in combination with the in gel GLDH activity assay and immunoblotting, two mitochondrial complexes of 470 and 420 kDa were identified. Both complexes are of very low abundance. Protein identifications by mass spectrometry revealed that they include subunits of complex I. Finally, the 850-kDa complex was further investigated and shown to include the complete "peripheral arm" of complex I. GLDH is attached to a membrane domain, which represents a major fragment of the "membrane arm" of complex I. Taken together, our data further support a role of GLDH during complex I formation, which is based on its binding to specific assembly intermediates

Impedance Control on Arbitrary Surfaces for Ultrasound Scanning using Discrete Differential Geometry

We propose an approach to robotic ultrasound scanning and interaction control with arbitrary surfaces using a passivity-based impedance control scheme. First, we introduce task coordinates depending on the geometry of the surface, which enable hands-on guidance of the robot along the surface, as well as teleoperated and autonomous ultrasound image acquisition. Our coordinates allow controlling the signed distance of the robot to the surface and alignment of the tool to the surface normal using classical impedance control. This corresponds to implicitly obtaining a foliation of parallel surfaces. By setting the desired signed distance negative, i.e., into the surface, we obtain passive contact forces and simultaneously provide an intuitive way to control the maximum penetration depth into the surface. We extend the approach to also incorporate coordinates allowing to control the specific point on the surface and, automatically, on all parallel surfaces. Finally, we demonstrate the performance of the controller on the seven degrees of freedom lightweight robot DLR Miro: the robot tracks complex trajectories while accurately keeping the desired distance to the surface and applying an almost constant contact force. Finally, we compare the approach to the state of the art

Task-Specific Robot Base Pose Optimization for Robot-Assisted Surgeries

Preoperative planning and intra-operative system setup are crucial steps to successfully integrate robotically assisted surgical systems (RASS) into the operating room. Efficiency in terms of setup planning directly affects the overall procedural costs and increases acceptance of RASS by surgeons and clinical personnel. Due to the kinematic limitations of RASS, selecting an optimal robot base location and surgery access point for the patient is essential to avoid potentially critical complications due to reachability issues. To this end, this work proposes a novel versatile method for RASS setup and planning based on robot capability maps (CMAPs). CMAPs are a common tool to perform workspace analysis in robotics, as they are in general applicable to any robot kinematics. However, CMAPs have not been completely exploited so far for RASS setup and planning. By adapting global CMAPs to surgical procedure-specific tasks and constraints, a novel RASS capability map (RASSCMAP) is generated. Furthermore, RASSCMAPs can be derived to also comply with kinematic access constraints such as access points in laparoscopy. RASSCMAPs are versatile and applicable to any kind of surgical procedure; they can be used on the one hand for aiding in intra-operative experience-based system setup by visualizing online the robot’s capability to perform a task. On the other hand, they can be used to find the optimal setup by applying a multi-objective optimization based on a genetic algorithm preoperatively, which is then transfered to the operating room during system setup. To illustrate these applications, the method is evaluated in two different use cases, namely, pedicle screw placement in vertebral fixation procedures and general laparoscopy. The proposed RASSCMAPs help in increasing the overall clinical value of RASS by reducing system setup time and guaranteeing proper robot reachability to successfully perform the intended surgerie