Lactose- and galactose-utilizing strains of poly(hydroxyalkanoic acid)-accumulating Alcaligenes eutrophus and Pseudomonas saccharophila obtained by recombinant DNA technology

Summary Transposon Tn 951-encoded -galactosidase was expressed in Pseudomonas saccharophila and enabled this bacterium to grow on lactose as sole carbon source. In contrast, -galactosidase was not expressed in Alcaligenes eutrophus even if the lacZ gene of Tn 951 was separated from the lacI gene. However, -galactosidase was expressed in A. eutrophus, if a DNA fragment, which was suspected to harbour the promoter of the A. eutrophus poly(3-hydroxybutyric acid)-synthetic genes, was ligated to the promoter probe vector pMC1403, which employs lac Z, Y as reporter genes. Plasmid pPL76, which harboured one of the promoter-lac fusions, enabled A. eutrophus not only to express -galactosidase but also to grow slowly on lactose (doubling time = 25–30 h). Subsequently, the promoter-lac fusion was ligated to Tn5 in pSUP5011 and was inserted into the genome of A. eutrophus H16 and of the glucose-utilizing mutant H16-G⁺1 by applying the suicide plasmid technique. Two recombinant strains, H16-cPL and H16-G⁺1-cPL, which grow with a doubling time of 16–23 h on lactose, were investigated in detail. The cells only utilized the glucose residue of lactose as a carbon source for grouth and excreted galactose into the medium. Only after the Escherichia coli gal operon had been cloned in vector pVK101 and had been mobilized to H16-cPL or H16-G⁺1-cPL, was lactose completely utilized; no galactose was detected in the medium and the growth yields increased twofold. Depending on the orientation of the gal operon in pVK101, the expression of galactokinase seems to be dependent either on the promoter of aminoglycoside phosphotransferase gene (kan) or on the promoter of the tetR gene. Offprint requests to: A. Steinbüchel     

Sea snake (Microcephalophis gracilis) hemoglobin: Primary structure and relationships to other forms

The hemoglobin of the sea snakeMicrocephalophis gracilis was purified and the primary structure of the α and β chains determined. This is the first sea snake hemoglobin structure characterized, and apparently also the first complete structure of any snake hemoglobin (an α chain of a viper was known), allowing judgments of reptilian variants. Variations between the sea snake form and other reptilian forms are large (52–65 differences for the α chains), of similar order as those between the sea snake and avian (56–65 differences) or human (58 differences) forms. Functionally, 19 residues at α/β contact areas and 7 at heme contacts are exchanged in relation to the human α and β chains. Four positions of the sea snake hemoglobin contain residues thus far unique to this form. However, all replacements appear compatible with conserved overall functional properties.     

Secretogranin II: Relative Amounts and Processing to Secretoneurin in Various Rat Tissues

Abstract: Secretoneurin is a 33-amino-acid peptide produced in vivo from secretogranin II. An antiserum raised against this peptide recognizes both the free peptide and its precursors. By HPLC and radioimmunoassay we characterized the immunoreactive molecules and determined the levels of immunoreactivity in various rat organs. In adrenal medulla and to a lesser degree in the anterior pituitary processing of secretogranin II to secretoneurin was very limited, whereas in all other organs studied (brain, intestine, endocrine pancreas, thyroid gland, and posterior pituitary) a high degree of processing was apparent. Thus, practically all of the immunoreactivity was present as free secretoneurin. This was also true for serum. When the total amount of secretoneurin immunoreactivity was calculated for the various organs, the largest pools in descending order were in the intestine, CNS, anterior pituitary, pancreas, and adrenal gland. This makes it likely that secretoneurin in serum is mainly derived from the intestine. The high degree of processing of secretogranin II in most organs is consistent with the concept that this protein acts as a precursor of a functional peptide, i.e., secretoneurin.     

Temperature influence on cohort parameters and demographic characteristics of the two cowpea coreids Clavigralla tomentosicollis and C. shadabi

Age-specific life tables of two important pests of cowpea, Vigna unguiculata (L.) Walp., the pod sucking bugs Clavigralla tomentosicollis Stål and C. shadabi Dolling (Heteroptera: Coreidae), were obtained from observations carried out at different temperatures. A biophysical model was found satisfactory to describe the temperature-response of developmental and mortality rates of egg and nymphal stages, with a peak developmental rate around 34°C in both species. The variability in development times was small and the experimental data did not permit any conclusion with regard to the Erlang probability density function. Survival of eggs and nymphs remained high between 20° and 30°C for both species. At temperatures above 34°C, C. tomentosicollis survivorship and fecundity was higher than that of C. shadabi, which in turn laid more eggs at temperatures between 20° and 30°C. Maximum fecundity is estimated to be at 29°C for C. tomentosicollis (99 eggs/female) and 26°C for C. shadabi (261 eggs/female). At 30°C, the intrinsic rate of increase reached a maximum in both species, 0.152 per day for C. tomentosicollis and 0.145 per day for C. shadabi, and remained high for C. tomentosicollis until 36°C. C. tomentosicollis performed significantly better on pigeonpea, Cajanus cajan Millsp., than on cowpea at higher temperatures.     

Langzeit-Erfassung brutbiologischer Parameter beim KleiberSitta europaea in zwei norddeutschen Untersuchungsr?umen

Zusammenfassung Der Zusammenstellung liegen Befunde zu den brutbiologischen Parametern Gelegestärke, Schlüpftermin und Anzahl flügger Junge zugrunde, die von 1970–1995 an rund 1500 Erstbruten des Kleibers in Versuchsgebieten mit künstlichen Nisthöhlen bei Braunschweig und Essen gewonnen wurden. Die Vollgelege-Eizahl beträgt im Durchschnitt 6,8 Eier (nur Daten aus dem Braunschweiger Raum), der durchschnittliche Schlüpftermin fällt auf den 9. Mai (Braunschweig) und 6. Mai (Essen) und die Anzahl flügger Nestlinge liegt in erfolgreichen Bruten im Mittel bei 5,6 (Braunschweig) bzw. 5,2 (Essen). Im Untersuchungszeitraum kam es in beiden Gebietsregionen zu einer signifikanten Verfrühung des Bruttermins. Doch werden wohl erst die Befunde der kommenden Jahre zeigen können, ob die rezente Häufung früher Brutjahre beim Kleiber (diese Arbeit) und bei Kohl- und Blaumeisen (Winkel &Hudde unveröff.) bereits eine Folge des anthropogen bedingten global warming ist oder noch das normale Auf und Ab der Witterung widerspiegelt.

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Long-term changes of breeding parameters of NuthatchesSitta europaea in two study areas of northern Germany

Between 1970 and 1995, nestbox data about clutch size, hatching date and fledging success were collected for a total of around 1500 first broods of Nuthatches in two north German study areas (near Brunswick 52.16 N 10.32 E and near Essen 51.24 N 6.59 E). The average clutch size was 6.8 eggs (Brunswick), the average hatching date 9 May (Brunswick) and 6 May (Essen), and the mean number of fledglings per successful brood 5.6 (Brunswick) and 5.2 (Essen). Over time hatching dates significantly advanced in both areas. Data of further years are needed to clarify if the recent unusual sequence of early breeding years in the Nuthatch is already a consequence of the anthropogenic greenhouse effect or just due to normal variation of weather conditions.

     

Reproductive success and clonal genetic structure of the rareArnica montana (Compositae) in The Netherlands

In a medium-sized population ofArnica montana, a threatened species in The Netherlands, the breeding system, reproductive success and genetic clonal structure were studied. Pollination experiments suggested thatA. montana is largely self-incompatible. Inbreeding depression was observed for seedling weight but not for fruit weight and germination rate. Although genetic variation is rather low in this population, the data suggest an outcrossing mating system. Analysis of the genotype of all mapped rosettes in a plot of 100 m² indicated that dense clusters often consist of identical genotypes, suggesting a clonal structure. Open clusters frequently contained several different genotypes. This may be caused by limited fruit dispersal, since seedlings were found mainly within or in the near surroundings of the clusters.     

High-conductance calcium-activated potassium channels; Structure,pharmacology, and function

High-conductance calcium-activated potassium (maxi-K) channels comprise a specialized family of K⁺ channels. They are unique in their dual requirement for depolarization and Ca²⁺ binding for transition to the open, or conducting, state. Ion conduction through maxi-K channels is blocked by a family of venom-derived peptides, such as charybdotoxin and iberiotoxin. These peptides have been used to study function and structure of maxi-K channels, to identify novel channel modulators, and to follow the purification of functional maxi-K channels from smooth muscle. The channel consists of two dissimilar subunits, and . The subunit is a member of theslo Ca²⁺-activated K⁺ channel gene family and forms the ion conduction pore. The subunit is a structurally unique, membrane-spanning protein that contributes to channel gating and pharmacology. Potent, selective maxi-K channel effectors (both agonists and blockers) of low molecular weight have been identified from natural product sources. These agents, together with peptidyl inhibitors and site-directed antibodies raised against and subunit sequences, can be used to anatomically map maxi-K channel expression, and to study the physiologic role of maxi-K channels in various tissues. One goal of such investigations is to determine whether maxi-K channels represent novel therapeutic targets.     

Effect of the redox state of QB on electric field-induced charge recombination in Photosystem II

Electric field-induced charge recombination in Photosystem II (PS II) was studied in osmotically swollen spinach chloroplasts (blebs) by measurement of the concomitant chlorophyll luminescence emission (electroluminescence). A pronounced dependence on the redox state of the two-electron gate Q_B was observed and the earlier failure to detect it is explained. The influence of the Q_B/Q_B ^– oscillation on electroluminescence was dependent on the redox state of the oxygen evolving complex; at times around one millisecond after flash illumination a large effect was observed in the states S₂ and S₃, but not in the state S₄ (actually Z⁺S₃). The presence of the oxidized secondary electron donor, tyrosine Z⁺, appeared to prevent expression of the Q_B/Q_B ^– effect on electroluminescence, possibly because this effect is primarily due to a shift of the redox equilibrium between Z/Z⁺ and the oxygen evolving complex.Abbreviations BSA bovine serum albumin - EDTA ethylene-diaminetetraacetic acid - EL electroluminescence - FCCP carbonylcyanide p-trifluoromethyloxyphenyl-hydrazone - HEPESI 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - I primary electron acceptor - MOPS 3-(N-morpholino) propane sulfonic acid - P680 primary electron donor of Photosystem II - P700 primary electron donor of Photosystem I - Q_A and Q_B secondary and tertiary electron acceptors of Photosystem II - Z secondary electron donor (D1 Tyr 161)     

Estimating denitrification rates in estuarine sediments: A comparison of stoichiometric and acetylene based methods

We compared denitrification rates obtained using an adaptation of the acetylene block technique to rates estimated from benthic flux nutrient stoichiometry in the subtidal sediments of Tomales Bay, California (USA). By amending whole cores with acetylene and saturating nitrate concentrations, we obtained potential denitrification rates, which ranged between 4 and 30 mmol N m^–2 d^–1. We determined the apparent Michaelis constant (K_app) and the maximum potential rate (V_mp) of the denitrifying community and used these constants in a rectangular hyperbola to estimatein situ denitrification rates. Both the K_app and V_mp of the denitrifying community exhibited significant variation over both depth in the sediment column and time of sampling.Estimates ofin situ denitrification obtained using our kinetic-fix adaptation of the acetylene block ranged between 1.8 (March) and 9 (Sept.) mmol N m^–1 d^–1. Denitrification rates obtained using benthic flux stoichiometry ranged between 0.7 and 4.1 mmol N m^–2 d^–1. Average denitrification rates obtained using the kinetic-fix acetylene block approach exceeded those obtained from net benthic flux stoichiometry; however, these differences were not significant. We conclude that our kinetic-fix adaptation of the acetylene block technique provides realistic estimates of denitrification in sediments, even when pore water nitrate concentrations are low and nitrification and denitrification are closely coupled.