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Bacillus subtilis citM, the structural gene for dihydrolipoamide transsuccinylase: cloning and expression in Escherichia coli

The 2-oxoglutarate dehydrogenase multienzyme complex is composed of three different subenzymes: 2-oxoglutarate dehydrogenase (E1o), dihydrolipoamide transsuccinylase (E2o), and dihydrolipoamide dehydrogenase (E3). Bacillus subtilis E1o and E2o are encoded by the citK and citM genes, respectively. A 3.4-kb BamHI DNA fragment containing citK and citM markers was isolated from a library of B. subtili

Genetic and biochemical characterization of Bacillus subtilis mutants defective in expression and function of cytochrome b-558

Bacillus subtilis succinate dehydrogenase is bound to the cytoplasmic membrane by cytochrome b-558, a 23-kDa transmembrane protein which also functions as electron acceptor to the dehydrogenase. The structural gene for the apocytochrome, sdhC, has previously been cloned and sequenced. In this work the structure and translation of cytochrome b-558 was studied in different sdhC mutants. Mutant cytoc

Processing of Bacillus subtilis succinate dehydrogenase and cytochrome b-558 polypeptides

The DNA sequence of the Bacillus subtilis sdh operon coding for the two succinate dehydrogenase subunits and cytochrome b-558 (the membrane anchor protein) has recently been established. We have now determined the extent of N-terminal processing of each polypeptide by radiosequence analysis. At the same time, direct evidence for the correctness of the predicted reading frames has been obtained. Th

Deletion of the Bacillus subtilis sdh operon

Plasmid pKIM2 carries the Bacillus subtilis sdh operon and adjacent regions of the bacterial chromosome. The plasmid replicates in Escherichia coli but not in B. subtilis. Different portions of the sdh operon were removed from pKIM2 and replaced by a cat gene derived from pC194. A series of plasmids carrying sdh deletions was thus derived. Plasmid DNA was linearized at restriction sites within the

Nucleotide sequence encoding the flavoprotein and the iron-sulfur protein subunits of the Bacillus subtilis PY79 succinate dehydrogenase complex

The nucleotide sequence of a 2.7-kilobase segment of DNA containing the sdhA and sdhB genes encoding the flavoprotein (Fp, sdhA) and iron-sulfur protein (Ip, sdhB) subunits of the succinate dehydrogenase of Bacillus subtilis was determined. This sequence extends the previously reported sequence encoding the cytochrome b558 subunit (sdhC) and completes the sequence of the sdh operon, sdhCAB. The pr

In vitro complementation of Bacillus subtilis and Escherichia coli 2-oxoglutarate dehydrogenase complex mutants and genetic mapping of B. subtilis citK and citM mutations

AbstractThe 2-oxoglutarate dehydrogenase complex of the tricarboxylic acid cycle (TCA) consists of multiple copies of 3 different subenzymes; E1, E2 and E3. The E3 subenzyme is also a component of the pyruvate dehydrogenase complex. Bacillus subtilis 2-oxoglutarate dehydrogenase mutants were studied. The mutants defective in E1, E2 and E3 subenzyme activity, respectively, could be separated into 3

Molecular properties, genetics, and biosynthesis of Bacillus subtilis succinate dehydrogenase complex

This chapter discusses the molecular properties, genetics, and biosynthesis of Bacillus subtilis succinate dehydrogenase complex. The citric acid cycle enzyme succinate dehydrogenase (SDH) is a membrane-bound iron-sulfur flavoprotein. Mitochondrial and bacterial SDH and membrane-bound fumarate reductase in anaerobic and facultative bacteria are similar in composition. SDH and also fumarate reducta

Rare Variant Analysis of Human and Rodent Obesity Genes in Individuals with Severe Childhood Obesity

Obesity is a genetically heterogeneous disorder. Using targeted and whole-exome sequencing, we studied 32 human and 87 rodent obesity genes in 2,548 severely obese children and 1,117 controls. We identified 52 variants contributing to obesity in 2% of cases including multiple novel variants in GNAS, which were sometimes found with accelerated growth rather than short stature as described previousl

Electron-paramagnetic-resonance spectroscopy of Bacillus subtilis cytochrome b558 in Escherichia coli membranes and in succinate dehydrogenase complex from Bacillus subtilis membranes

Cytochrome b558 of the Bacillus subtilis succinate dehydrogenase complex was studied by electron-paramagnetic-resonance (EPR) spectroscopy. The cytochrome amplified in Escherichia coli membranes by expression of the cloned cytochrome gene and in the succinate dehydrogenase complex immunoprecipitated from solubilized B. subtilis membranes, respectively, is shown to be low spin with a highly anisotr

Modification of bovine heart succinate dehydrogenase with ethoxyformic anhydride and Rose bengal: Evidence for essential histidyl residues protectable by substrates

Purified and membrane-bound succinate dehydrogenase (SDH) from bovine heart mitochondria was inhibited by the histidine-modifying reagents ethoxyformic anhydride (EFA) and Rose Bengal in the presence of light. Succinate and competitive inhibitors protected against inhibition, and decreased the number of histidyl residues modified by EFA. The essential residue modified by EFA was not the essential

Bacillus subtilis mutant succinate dehydrogenase lacking covalently bound flavin: Identification of the primary defect and studies on the iron-sulfur clusters in mutated and wild- type enzyme

Succinate dehydrogenase consists of two protein subunits and contains one FAD and three iron-sulfur clusters. The flavin is covalently bound to a histidine in the larger, Fp, subunit. The reduction oxidation midpoint potentials of the clusters designated S-l, S-2, and S-3 in Bacillus subtilis wild-type membrane-bound enzyme were determined as +80, -240, and -25 mV, respectively. Magnetic spin inte

Effect of burner geometry on swirl stabilized methane/air flames : A joint LES/OH-PLIF/PIV study

Large eddy simulation (LES) using a transported PDF model and OH-PLIF/PIV experiments were carried out to investigate the quarl effects on the structures of swirl stabilized methane/air flames. Two different quarls were investigated, one straight cylindrical quarl and one diverging conical quarl. The experiments show that the flames are significantly different with the two quarls. With the straigh

Charging Information Services for BEVs – Two Competing and Complementary Business Models

This paper examines information services in association with charging services, and in particular players that offer information services related to charging service offerings from many different operators. It identifies two main types of business models through studying and comparing 25 European charging information service providers. For one type of players the paper analyses the network charact

Influence of prefiltration on membrane performance during isolation of lignin-carbohydrate complexes from spent sulfite liquor

In this study, we examined the isolation of lignin-carbohydrate complexes (LCCs) from sodium-based spent sulfite liquor, in conjunction with minimization of membrane fouling. We screened 3 polysulfone (PS) membranes with cutoffs of 100, 50, and 25 kDa, respectively. Flux and retention for the 100- and 50-kDa membranes had the same order of magnitude, indicating that these properties were determine

Lactose, glucose and galactose content in milk, fermented milk and lactose-free milk products

Lactose, glucose and galactose contents in milk and fermented milk products and their lactose-free alternatives, were determined. Storage of products up to the best-before dates had no effect on carbohydrate composition. Total galactose content in yoghurt, i.e., free galactose plus the galactose moiety of lactose, was 2.3–2.4 g 100 g−1 (94–95% of that in milk), whereas that in Swedish soured milk

Flow rate analysis of compressible superheated steam through pressure reducing valves

Steam flow is the main form for energy transfer in power plants, process industries, etc. The flow rate of the steam relates to the energy transfer amount directly. Pressure reducing valves are used for flow rate control of the compressible superheated steam in these piping systems. In this paper, a pressure reducing valve with a novel valve core is proposed. In order to analyze the internal flow

Pressure Drop Analysis of Pilot-Control Globe Valve With Different Structural Parameters

Pilot-control globe valve (PCGV) can use the pressure drop caused by fluid flowing through the orifice located at valve core bottom to open or close the main valve using a small pilot valve. In this paper, computational fluid dynamics (CFD) method is adopted to analyze the pressure drop before and after valve core of PCGV and minor loss of orifice under different structural parameters and inlet ve

Radiocarbon calibration uncertainties during the last deglaciation : Insights from new floating tree-ring chronologies

Radiocarbon dating is the most commonly used chronological tool in archaeological and environmental sciences dealing with the past 50,000 years, making the radiocarbon calibration curve one of the most important records in paleosciences. For the past 12,560 years, the radiocarbon calibration curve is constrained by high quality tree-ring data. Prior to this, however, its uncertainties increase rap

Ecosystem model analysis of multi-use forestry in a changing climate

The challenge of multi-use forestry is to fulfil a range of economic, ecologic and social goals in a sustainable way, accounting for synergies and trade-offs among the ecosystem services provided. Climate changes add to the complexity via effects on forest ecosystem processes, such as primary production and respiration, and also by adding a new goal on the agenda: the role of forests in climate mi

Data on haplotype-supported immunoglobulin germline gene inference

Data that defines IGHV (immunoglobulin heavy chain variable) germline gene inference using sequences of IgM-encoding transcriptomes obtained by Illumina MiSeq sequencing technology are described. Such inference is used to establish personalized germline gene sets for in-depth antibody repertoire studies and to detect new antibody germline genes from widely available immunoglobulin-encoding transcr