The antenna cleaner gland in Messor rufitarsis (hymenoptera, formicidae)

TEM and SEM analysis of the tibio-tarsal antennal cleaner in Messor rufitarsis reveals a pore region on the surface of the basitarsus. These pores are originating from a cluster of highly prismatic epidermal cells that fill the anterior hemolymph space of the leg almost completely. Within the cytoplasma of the cells, basal labyrinth, nucleus/rough endoplasmic reticulum, 'secondary lysosomes', a lot of smooth endoplasmic reticulum and distal microvilli regions can be distinguished indicating that the cells are 'class I' gland cells. Their secretion appears to be transported to the antennal cleaner apparatus via a subcuticular space and pores. It is suggested that the secretion might be used for cleaning of the antenna and/or chemical communication. SEM examination of six additional species indicates that an antennal cleaner gland might be present throughout the ants.     

Sperm viability is influenced in vitro by the bovine seminal protein aSFP: effects on motility, mitochondrial activity and lipid peroxidation

The 13 kDa acidic seminal fluid protein (aSFP) is a major component of bovine semen exerting growth factor-like activity. The influence of the pure protein on sperm viability was observed by evaluating sperm motility using computer-assisted semen analysis. Furthermore, mitochondrial dehydrogenase activity as a parameter of sperm metabolism and the integrity of sperm membranes using a metal catalyzed lipid peroxidation assay were measured. Over a wide physiological range (0.003 to 4 g/l) aSFP did not influence motility and average-path velocity of sperm, but at the highest concentration (6 g/l) a significant reduction in motility could be observed. Mitochondrial activity was significantly stimulated at medium concentrations (0.125 to 2 g/l), whereas a 40% suppression was observed at maximum levels (4 g/l). A dose-dependent inhibition of lipid peroxidation could be demonstrated for medium and high concentrations of aSFP (0.125 to 4 g/l). Compared with other reducing agents, aSFP showed the highest potency in preventing oxidative stress. Such effects might be explained by the remarkable redox behavior of the protein. We suggest that in the bull aSFP may play a role in the regulation of sperm metabolism and the protection of sperm membranes from oxidative damage.     

Electrooptical analysis of alpha-chymotrypsin at physiological salt concentration

The electric dichroism of alpha-chymotrypsin has been measured in a buffer containing 0.1 M Na(+), 10 mM Mg(2+) and 25 mM Tris-cacodylate pH 7.2. The reduced dichroism as a function of the electric field strength can be represented by the orientation function for permanent dipoles and is not consistent with the orientation function for induced dipoles. After correction for the internal directing field, the dipole moment is 1.1 x 10(-27) Cm (+/- 10%), corresponding to 340 D, at 20 degrees C. The assignment of the permanent dipole moment is confirmed by the shape of the dichroism rise curves, which require two exponentials with amplitudes of opposite sign for fitting. The dichroism decay time constants measured in the range of temperatures between 2 and 30 degrees C indicate a temperature induced change of the structure, which is equivalent to an increase of the hydrodynamic radius from r = 26.6 A at 2 degrees C to 28.5 A at 30 degrees C. Our results demonstrate that electrooptical investigations of proteins with a high time resolution can be extended to physiological salt concentrations without serious problems by use of appropriate instruments.     

Kinetics of excited states of pigment clusters in solubilized light-harvesting complex II: photon density-dependent fluorescence yield and transmittance.

Relative fluorescence yield, phi F, and transmittance, T, were measured in solubilized light-harvesting complex II (LHCII) as a function of photon density, Ip, of monochromatic 645-nm laser pulses (duration: approximately 2.5 ns). Special efforts were made in constructing an optical set-up that allows the accurate determination of the fluorescence from an area of constant Ip, phi F(Ip) starts to decline at approximately 10(14) and drops to values below 0.01% at maximum Ip (approximately 10(19) photons cm-2 pulse-1). T(Ip) decreases only slightly at photon densities of approximately 10(15) but increases steeply at values of > 10(17) photons cm-2 pulse-1. The interpretation of the phi F(Ip) data using the saturation limit of Mauzerall's multiple hit model leads to a unit size of about 10-15 chlorophyll molecules. One interpretation is to attribute this result to a very fast exciton-exciton annihilation of multiple excited states generated within this small domain. Alternatively, based on the assumption that delocalized cluster states within the monomeric/trimeric subunit of LHCII exist, the results can be consistently described by a kinetic model comprising ground, monoexcitonic, and biexcitonic states of clusters and a triplet state that is quenched by carotenoids in LHCII. Within the framework of this model the annihilation of multiple excitations is explained as ultrafast radiationless relaxation of higher excited cluster states. Comparative measurements in diluted acetonic Chl a solution are consistently described by the depletion of the ground state, taking the absorption cross section at the used wavelength.     

Hybridomas in a bioreactor cascade: modeling and determination of growth and death kinetics

Hybridomas were cultured under steady-state conditions in a series of two continuous stirred-tank reactors (CSTRs), using a serum-free medium. The substrate not completely converted in the first CSTR, was transported with the cells to the second one and very low growth rates, high death rates, and lysis of viable cells were observed in this second CSTR. These conditions are hardly accessible in a single vessel, because such experiments would be extremely time-consuming and unstable due to a low viability. In contrast to what is often observed in literature, kinetic parameters could thus be derived without the neccessity for extrapolation to lower growth rates. Good agreement with literature averages for other hybridomas was found. Furthermore, showing that the reactor series is a valuable research tool for kinetic studies under extreme conditions, the possibility to observe cell death under stable and defined steady-state conditions offers interesting opportunities to investigate apoptosis and necrosis. Additionally, a model was developed that describes hybridoma growth and monoclonal antibody production in the bioreactor cascade on the basis of glutamine metabolism. Good agreement between the model and the experiments was found.Abbreviation MAb Monoclonal antibody Nomenclature C _AConcentration of any (mol m^-3) component A D Dilution rate (s^-1) K _dDeath-rate constant (mol m^-3) K _lLysis-rate constant (mol m^-3) K _sMonod constant (mol m^-3) m Maintenance coefficient (mol cell^-1 s^-1) q Specific consumption (mol cell^-1 s^-1) or production rate t Time (s) X Cell concentration (cell m^-3) Y Yield coefficient (cell mol^-1) Greek symbols _d Specific death rate (s^-1) _l Specific lysis rate (s^-1) of viable cells _net Net specific growth (s^-1) rate _true True specific growth (s^-1) rate     

Biodegradation of 4-nitroanisole by two Rhodococcus spp.

Two Rhodococcus strains, R. opacus strain AS2 and R. erythropolis strain AS3, that were able to use 4-nitroanisole as the sole source of carbon and energy, were isolated from environmental samples. The first step of the degradation involved the O-demethylation of 4-nitroanisole to 4-nitrophenol which accumulated transiently in the medium during growth. Oxygen uptake experiments indicated the transformation of 4-nitrophenol to 4-nitrocatechol and 1,2,4-trihydroxybenzene prior to ring cleavage and then subsequent mineralization. The nitro group was removed as nitrite, which accumulated in the medium in stoichiometric amounts. In R. opacus strain AS2 small amounts of hydroquinone were produced by a side reaction, but were not further degraded.     

Cloning, expression, and chromosomal localization of the 140-kilodalton subunit of replication factor C from mice and humans.

We have isolated a full-length mouse cDNA encoding a lysine-rich protein of 1,131 amino acids with a calculated molecular mass of 126 kDa. The protein binds in a sequence-unspecific manner to DNA, is localized exclusively in the nucleus, and contains a putative ATP binding site and a stretch of 80 amino acids with homology to the carboxy terminus of prokaryotic DNA ligases. On the basis of the following facts, we conclude that the isolated cDNA encodes the 140-kDa subunit of mouse replication factor C (mRFC140). (i) The sequence around the ATP binding site shows significant homology to three small subunits of human replication factor C. (ii) Polyclonal antibodies raised against the protein encoded by this cDNA cross-react with the 140-kDa subunit of purified human replication factor C (hRFC140) and recognize in mouse cell extracts an authentic protein with an apparent molecular mass of 130 kDa. (iii) Sequence comparison with a human cDNA isolated by using tryptic peptide sequence information from purified hRFC140 revealed 83% identity of the encoded proteins. The mRFC140 gene is ubiquitously expressed, and two mRNAs approximately 5.0 and 4.5 kb long have been detected. The gene was mapped by in situ hybridization to mouse chromosome 5, and its human homolog was mapped to chromosome 4 (p13-p14).     

Bicyclic carbo- and heterocycles from an allylidene complex and cyclic 1,3-dienes by tandem cyclopropanation/cope rearrangement

The allylidene complex (CO)₅W=CH---C(Ph)=C(Ph)H (4) reacts with cyclopentadiene by stereospecific transfer of the carbene ligand to one of the two double bonds of cyclopentadiene to give a cis-divinylcyclopropane complex 5. The divinylcyclopropane ligand coordinates to the metal via the unsubstituted double bond. Addition of bromide to solutions of 5 gives rise to the formation of [(CO)₅WBr]⁻ and a bicyclo[3.2.1]octadiene (6), the Cope rearrangement product of the free divinylcyclopropane. Thermolysis of 5 affords 6 and its (CO)₅W complex. The reaction of 4 with furan (8a), 2-methylfuran (8b) and 3-methylfuran (8c) affords the (CO)₅W(bicyclo[3.2.1]oxahepta- diene) complexes (9a–c), The formation of 9a–c which is chemo-, regio- and stereospecific is explained by a tandem cyclopropanation/Cope rearrangement sequence. The bicyclic ligands 10a–c are liberated from the metal either by thermolysis of solutions of 9a–c or by addition of bromide.     

α2-Adrenoceptor-Mediated Inhibition of Electrically Evoked [3H]Noradrenaline Release from Chick Sympathetic Neurons: Role of Cyclic AMP

Abstract: This study explores the role of cyclic AMP in electrically evoked [³H]noradrenaline release and in the α₂-adrenergic modulation of this release in chick sympathetic neurons. Along with an increase in stimulation-evoked tritium overflow, applications of forskolin enhanced the formation of intracellular cyclic AMP. Both effects of forskolin were potentiated by the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine. The forskolin-induced increase in overflow was abolished by the Rp-diastereomer of cyclic AMP-thioate, an antagonist at cyclic AMP-dependent protein kinases, and 1,9-dideoxy-forskolin, an inactive analogue at adenylyl cyclase, had no effect on the evoked overflow. A 24-h pretreatment with either cholera toxin or forskolin reduced the subsequent forskolin-induced accumulation of cyclic AMP and inhibited the stimulation-evoked release. Basal cyclic AMP production, however, remained unaltered after forskolin treatment and was enhanced after 24 h of cholera toxin exposure. The α₂-adrenergic agonist bromoxidine did not affect the formation of cyclic AMP stimulated by forskolin but reduced electrically evoked release. However, effects of bromoxidine on ³H overflow were attenuated by forskolin as well as by 8-bromo-cyclic AMP. Effects of bromoxidine on [³H]noradrenaline release were paralleled by an inhibition of voltage-activated Ca²⁺ currents, primarily through a delayed time course of current activation. This effect was abolished when either forskolin or 8-bromo-cyclic AMP was included in the pipette solution. Both substances, however, failed to affect Ca²⁺ currents in the absence of bromoxidine. These results suggest that the signaling cascade of the α₂-adrenergic inhibition of noradrenaline release involves voltage-activated Ca²⁺ channels but not cyclic AMP. Elevated levels of cyclic AMP, however, antagonize this α₂-adrenergic reduction, apparently through a disinhibition of Ca²⁺ channels.