Archaeal complex II: 'classical' and 'non-classical' succinate:quinone reductases with unusual features.

Reversible succinate dehydrogenase (SDH) activities have been ubiquitously detected in organisms from the three domains of life. They represent constituents either of respiratory complexes II in aerobes, or of fumarate dehydrogenase complexes in anaerobes. The present review gives a survey on archaeal succinate:quinone oxidoreductases (SQRs) analyzed so far. Though some of these could be studied in detail enzymologically and spectroscopically, the existence of others has been deduced only from published genome sequences. Interestingly, two groups of enzyme complexes can be distinguished in Archaea. One group resembles the properties of SDHs known from bacteria and mitochondria. The other represents a novel class with an unusual iron-sulfur cluster in subunit B and atypical sequence motifs in subunit C which may influence electron transport mechanisms and pathways. This novel class of SQRs is discussed in comparison to the so-called 'classical' complexes. A phylogenetic analysis is presented suggesting a co-evolution of the flavoprotein-binding subunit A and subunit B containing the three iron-sulfur clusters.     

Aminergic neuron systems of lobsters: morphology and electrophysiology of octopamine-containing neurosecretory cells

In the American lobster (Homarus americanus) the biogenic amines serotonin and octopamine appear to play important and opposite roles in the regulation of aggressive behavior, in the establishment and/or maintenance of dominant and subordinate behavioral states and in the modulation of the associated postural stances and escape responses. The octopamine-containing neurosecretory neurons in the thoracic regions of the lobster ventral nerve cord fall into two morphological subgroups, the root octopamine cells, a classical neurohemal group with release regions along second thoracic roots, and the claw octopamine cells, a group that selectively innervates the claws. Cells of both subgroups have additional sets of endings within neuropil regions of ganglia of the ventral nerve cord. Octopamine neurosecretory neurons generally are silent, but when spontaneously active or when activated, they show large overshooting action potentials with prominent after-hyperpolarizations. Autoinhibition after high-frequency firing, which is also seen in other crustacean neurosecretory cells, is readily apparent in these cells. The cells show no spontaneous synaptic activity, but appear to be excited by a unitary source. Stimulation of lateral or medial giant axons, which excite serotonergic cells yielded no response in octopaminergic neurosecretory cells and no evidence for direct interactions between pairs of octopamine neurons, or between the octopaminergic and the serotonergic sets of neurosecretory neurons was found.     

Ontogeny and homology of the skeletal elements that form the sucking disc of remoras (Teleostei,Echeneoidei, Echeneidae)

The sucking disc of the sharksuckers of the family Echeneidae is one of the most remarkable and most highly modified skeletal structures among vertebrates. We studied the development of the sucking disc based on a series of larval, juvenile, and adult echeneids ranging from 9.3 mm to 175 mm standard length. We revisited the question of the homology of the different skeletal parts that form the disc using an ontogenetic approach. We compared the initial stages of development of the disc with early developmental stages of the spinous dorsal fin in a representative of the morphologically basal percomorph Morone. We demonstrate that the “interneural rays” of echeneids are homologous with the proximal‐middle radials of Morone and other teleosts and that the “intercalary bones” of sharksuckers are homologous with the distal radials of Morone and other teleosts. The “intercalary bones” or distal radials develop a pair of large wing‐like lateral extensions in echeneids, not present in this form in any other teleost. Finally the “pectinated lamellae” are homologous with the fin spines of Morone and other acanthomorphs. The main part of each pectinated lamella is formed by bilateral extensions of the base of the fin spine just above its proximal tip, each of which develops a row of spinous projections, or spinules, along its posterior margin. The number of rows and the number of spinules increase with size, and they become autogenous from the body of the lamellae. We also provide a historical review of previous studies on the homology of the echeneid sucking disc and demonstrate that the most recent hypotheses, published in 2002, 2005 and 2006, are erroneous. J. Morphol. 2012. © 2012 Wiley Periodicals, Inc.     

A familial mutation in the testis-determining gene SRY shared by both sexes

A familial mutation in SRY, the gene coding for the testis-determining factor TDF, was identified in an XY female with gonadal dysgenesis, her father, her two brothers and her uncle. The mutation consists of a T to C transition in the region of the SRY gene coding for a protein motif known as the high mobility group (HMG) box, a protein domain known to confer DNA-binding specificity on the SRY protein. This point mutation results in the substitution, at amino acid position 109, of a serine residue for phenylalanine, a conserved aromatic residue in almost all HMG box motifs known. This F109S mutation was not found in 176 male controls. When recombinant wildtype SRY and SRY^F109S mutant protein were tested in vitro for binding to the target site AAC AAAG, no differences in DNA-binding activity were observed. These results imply that the F109S mutation either is a rare neutral sequence variant, or produces an SRY protein with slightly altered in vivo activity, the resulting sex phenotype depending on the genetic back-ground or environmental factors.This paper is dedicated by G. S. to Professor Ulrich Wolf on the occasion of his 60th birthday     

Metabolism of nitric oxide and nitrous oxide during nitrification and denitrification in soil at different incubation conditions

Abstract NO production and consumption rates as well as N₂O accumulation rates were measured in a loamy cambisol which was incubated under different conditions (i.e. soil moisture content, addition of nitrogen fertilizer and/or glucose, aerobic or anaerobic gas phase). Inhibition of nitrification with acetylene allowed us to distinguish between nitrification and denitrification as sources of NO and N₂O. Under aerobic conditions untreated soil showed very low release of NO and N₂O but high consumption of NO. Fertilization with NH₄⁺ or urea stimulated both NO and N₂O production by nitrification. Addition of glucose at high soil moisture contents led to increased N₂ and N₂O production by denitrification, but not to increased NO production rates. Anaerobic conditions, however, stimulated both NO and N₂O production by denitrification. The production of NO and N₂O was further stimulated at low moisture contents and after addition of glucose or NO₃⁻. Anaerobic consumption of NO by denitrification followed Michaelis-Menten kinetics and was stimulated by addition of glucose and NO₃⁻. Aerobic consumption of NO followed first-order kinetics up to mixing ratios of at least 14 ppmv NO, was inhibited by autoclaving but not by acetylene, and decreased with increasing soil moisture content. The high NO-consumption activity and the effects of soil moisture on the apparent rates of anaerobic and aerobic production and consumption of NO suggest that diffusional constraints have an important influence on the release of NO, and may be a reason for the different behaviour of NO release vs N₂O release.     

The membrane dialysis bioreactor with integrated radial-flow fixed bed —a new approach for continuous cultivation of animal cells

A hybridoma cell was cultivated continuously in a membrane dialysis bioreactor with an integrated radial-flow fixed bed consisting of porous Siran^® carriers over a period of 6 weeks. Antibodies accumulated to an average of 100 mg l^?1, approx. 10 times more than in fixed bed cultures without dialysis membrane. Serum costs could be reduced about 85% due to an appropriate feeding strategy. Siran^® carriers with 3–5 mm diameter showed an advantage compared to those with 1–2 mm diameter. For the 3–5 mm carrier the specific glucose uptake rate and the MAb production rate were constant, if the velocity was between 0.09 mm s^?1 and 0.75 mm s^?1. At higher velocities cells are washed out of the bed. Furthermore antibody consistency and cell stability were verified in long-term cultivations over a period of 96 days. From an estimation of the antibody concentration reachable with the reactor concept under optimal conditions a concentration 45 times higher compared to axial-flow fixed bed reactors and 11 times higher compared to stirred tank reactors can be expected.     

DNA polymerase alpha-DNA primase from human lymphoblasts

The DNA polymerase alpha-DNA primase complex from the human lymphoblast line HSC93 has been enriched to near homogeneity by using an immunoaffinity purification protocol which was developed earlier for the purification of the calf thymus enzyme (Nasheuer, H.-P. and Grosse, F. (1987) Biochemistry 26, 8458-8466). Immunoaffinity purified polymerase-primase from human cells consisted of four subunits displaying molecular weights of 195,000 and 180,000 for the DNA synthesizing alpha-subunit, of 68,000 for the beta-subunit, and of 55,000 and 48,000 for the primase-carrying gamma- and delta-subunit, respectively. The isoelectric pH values for the individual subunits were estimated from non-equilibrium pH gradients to be between 5.9 and 5.7 for the alpha-subunit, at 5.5 for the beta-subunit, and at 7.5 and 8.0 for the gamma- and delta-subunit, respectively. The purified polymerase-primase converted single-stranded phi X174 DNA into the double-stranded form in a primase-initiated reaction. During this process, 3-10 RNA primers were formed. RNA primers were about 11 nucleotides long. Elongation of existing RNA primers by the human polymerase-primase was semi-processive; following primer binding the DNA polymerase continuously incorporated 20 to 50 nucleotides, then it dissociated from the template DNA.     

Isolation and Partial Characterization of the Major Amide-Linked Conjugate of Indole-3-Acetic Acid from Phaseolus vulgaris L

A major indole-3-acetic acid conjugate from Phaseolus vulgaris seed has been isolated and partially characterized. It is a 3 kilodalton peptide with apparently 2 indole-3-acetyl moieties in amide linkage per peptide. The indole-3-acetic acid component was identified by gas chromatography-mass spectrometry and the peptide characterized by polyacrylamide gel electrophoresis, by amino acid analysis using dabsyl derivatives and by its Fourier transform-infrared spectrum. This is the first higher molecular weight amide-linked indole-3-acetic acid conjugate to be characterized from higher plants.     

The amount and nature of glutathione transferases in rat liver microsomes determined by immunochemical methods

The amount and nature of glutathione transferases in rat liver microsomes were determined using immunological techniques. It was shown that cytosolic glutathione transferase subunits A plus C, and B plus L were present at levels of 2.4 ± 0.6 and 1.5 ± 0.1 μg/mg microsomal protein, respectively. These levels are 10-times higher than those for non-specific binding of cytosolic components judging from the distribution of lactate dehydrogenase, a cytosolic marker. The possibility that a portion of these glutathione transferases is functionally localized on the endoplasmic reticulum is discussed. A previously described microsomal glutathione transferase which is distinct from the cytosolic enzymes is present in an amount of 31 ± 6 μg/mg microsomal protein.