First Measurement Using Elliptically Polarized Photons of the Double-Polarization Observable E for γp→pπ

We report the measurement of the helicity asymmetry E for the pπ^{0} and nπ^{+} final states using, for the first time, an elliptically polarized photon beam in combination with a longitudinally polarized target at the Crystal Ball experiment at MAMI. The results agree very well with data that were taken with a circularly polarized photon beam, showing that it is possible to simultaneously measure polarization observables that require linearly (e.g., G) and circularly polarized photons (e.g., E) and a longitudinally polarized target. The new data cover a photon energy range 270-1400 MeV for the pπ^{0} final state (230-842 MeV for the nπ^{+} final state) and the full range of pion polar angles, θ, providing the most precise measurement of the observable E. A moment analysis gives a clear observation of the pη cusp in the pπ^{0} final state.

Measurement of polarization observables T, P, and H in π0 and η photoproduction off quasi-free nucleons.

The target asymmetry T, recoil asymmetry P, and beam-target double polarization observable H were determined in exclusive π0 and η photoproduction off quasi-free protons and, for the first time, off quasi-free neutrons. The experiment was performed at the electron stretcher accelerator ELSA in Bonn, Germany, with the Crystal Barrel/TAPS detector setup, using a linearly polarized photon beam and a transversely polarized deuterated butanol target. Effects from the Fermi motion of the nucleons within deuterium were removed by a full kinematic reconstruction of the final state invariant mass. A comparison of the data obtained on the proton and on the neutron provides new insight into the isospin structure of the electromagnetic excitation of the nucleon. Earlier measurements of polarization observables in the γp→π0p and γp→ηp reactions are confirmed. The data obtained on the neutron are of particular relevance for clarifying the origin of the narrow structure in the ηn system at W=1.68GeV. A comparison with recent partial wave analyses favors the interpretation of this structure as arising from interference of the S11(1535) and S11(1650) resonances within the S11-partial wave.

Measurement of Compton Scattering at MAMI for the Extraction of the Electric and Magnetic Polarizabilities of the Proton.

A precise measurement of the differential cross sections dσ/dΩ and the linearly polarized photon beam asymmetry Σ_{3} for Compton scattering on the proton below pion threshold has been performed with a tagged photon beam and almost 4π detector at the Mainz Microtron. The incident photons were produced by the recently upgraded Glasgow-Mainz photon tagging facility and impinged on a cryogenic liquid hydrogen target, with the scattered photons detected in the Crystal Ball/TAPS setup. Using the highest statistics Compton scattering data ever measured on the proton along with two effective field theories (both covariant baryon and heavy-baryon) and one fixed-t dispersion relation model, constraining the fits with the Baldin sum rule, we have obtained the proton electric and magnetic polarizabilities with unprecedented precision: α_{E1}=10.99±0.16±0.47±0.17±0.34, ß_{M1}=3.14±0.21±0.24±0.20±0.35; in units of 10^{-4} fm^{3} where the errors are statistical, systematic, spin polarizability dependent, and model dependent.

Single π 0 production off neutrons bound in deuteron with linearly polarized photons.

The quasifree γ → d → π 0 n ( p ) photon beam asymmetry, Σ , has been measured at photon energies, E γ , from 390 to 610 MeV, corresponding to center of mass energy from 1.271 to 1.424 GeV, for the first time. The data were collected in the A2 hall of the MAMI electron beam facility with the Crystal Ball and TAPS calorimeters covering pion center-of-mass angles from 49 ∘ to 148 ∘ . In this kinematic region, polarization observables are sensitive to contributions from the Δ ( 1232 ) and N(1440) resonances. The extracted values of Σ have been compared to predictions based on partial-wave analyses (PWAs) of the existing pion photoproduction database. Our comparison includes the SAID, MAID and Bonn-Gatchina analyses; while a revised SAID fit, including the new Σ measurements, has also been performed. In addition, isospin symmetry is examined as a way to predict π 0 n photoproduction observables, based on fits to published data in the channels π 0 p , π + n and π - p .

Helicity-Dependent Cross Sections for the Photoproduction of π

The double-polarization observable E and helicity-dependent cross sections σ_{1/2}, σ_{3/2} have been measured for the photoproduction of π^{0} pairs off quasifree protons and neutrons at the Mainz MAMI accelerator with the Crystal Ball/TAPS setup. A circularly polarized photon beam was produced by bremsstrahlung from longitudinally polarized electrons and impinged on a longitudinally polarized deuterated butanol target. The reaction products were detected with an almost 4π covering calorimeter. The results reveal for the first time the helicity- and isospin-dependent structure of the γN→Nπ^{0}π^{0} reaction. They are compared to predictions from reaction models in view of nucleon resonance contributions and also to a refit of one model that predicted results for the proton and for the neutron target. The comparison of the prediction and the refit demonstrates the large impact of the new data.

Signatures of the d

We report a measurement of the spin polarization of the recoiling neutron in deuterium photodisintegration, utilizing a new large acceptance polarimeter within the Crystal Ball at MAMI. The measured photon energy range of 300-700 MeV provides the first measurement of recoil neutron polarization at photon energies where the quark substructure of the deuteron plays a role, thereby providing important new constraints on photodisintegration mechanisms. A very high neutron polarization in a narrow structure centered around E_{γ}∼570 MeV is observed, which is inconsistent with current theoretical predictions employing nucleon resonance degrees of freedom. A Legendre polynomial decomposition suggests this behavior could be related to the excitation of the d^{*}(2380) hexaquark.

Microbial degradation of halogenated compounds.

The mode of degradation of various halogenated compounds in isolated pure cultures and the disposition of the degradative genes have been studied. In many cases the degradative genes are found to be clustered on plasmids and appear to be under positive control. Genetic selection in vivo and genetic manipulations in vitro have allowed construction of strains having wider biodegradative potentials than their natural counterparts. Molecular cloning of the degradative gene clusters for halogenated compounds in vectors with a broad host range also allows the transfer of such genes to a large number of Gram-negative bacteria. The application of 2,4,5-trichlorophenoxyacetic acid (2,4,5-T)-degrading microorganisms has demonstrated the effectiveness of this strain in removing large amounts of 2,4,5-T from contaminated soil within a short period, and such soil has been shown to support the growth of plants normally sensitive to low concentrations of 2,4,5-T. The two major challenges that must be addressed in the near future are the development of appropriate microbial technology for the decontamination of soil containing hazardous halogenated compounds, and the promulgation of appropriate regulations to ensure the safety and well-being of the public during the application of genetically improved strains in an open environment.

Endoplasmic reticulum lumenal proteins of rat mammary gland. Potential involvement in lipid droplet assembly during lactation.

Intracellular distribution of selected reticuloplasmins, soluble proteins of the endoplasmic reticulum lumen, in rat mammary gland was investigated during pregnancy, lactation, and involution. During lactation the levels of the calcium binding protein calreticulin, and of protein disulfide isomerase, were elevated. Endoplasmic reticulum was as efficient as Golgi apparatus in sequestration and accumulation of Ca2+ from surrounding medium, as suggested from in vitro experiments with isolated cell fractions. Both protein disulfide isomerase and calreticulin were present in cytosol from homogenates of mammary gland prepared under mild conditions. Protein disulfide isomerase was abundant in intracellular lipid droplet precursors of milk lipid globules. Calreticulin and immunoglobulin binding protein (BiP, GRP 78) were associated with lipid droplets. Glucose-regulated protein (GRP 94) was not detected in association with intracellular lipid droplets. Milk lipid globule membrane lacked more than barely detectable quantities of protein disulfide isomerase, calreticulin, and immunoglobulin binding protein, suggesting that these proteins are lost from intracellular lipid droplets before or during their secretion as milk lipid globules. Immunocytochemical localization confirmed the presence of protein disulfide isomerase or calreticulin on intracellular lipid droplets and in non-endoplasmic reticulum regions of cells.

Low molecular mass GTP-binding proteins are secreted from mammary epithelial cells in association with lipid globules.

Secretion of milk lipid globules is achieved through encapsulation of triacylglycerol-rich lipid droplets in a specialized region of apical plasma membrane of mammary epithelial cells. A class of low molecular mass GTP-binding proteins were associated tightly with the lipid globule membrane, and these proteins appeared to change from peripheral to integral membrane proteins during intracellular growth and transit of lipid globule precursors. Inclusion of GTP or GTP gamma S in incubation medium stimulated secretion of lipids from primary cultures of permeabilized rat mammary epithelial cells. Six polypeptides with molecular masses between 28 and 21 kDa were detected by ability to bind GTP gamma S following separation of lipid-globule-associated proteins by SDS-PAGE and transblotting onto nitrocellulose. That all of these polypeptides were distinct immunologically from the archetype ras was evident from lack of immunoreactivity with p21 ras G-protein monoclonal antibody in Western blots. This monoclonal antibody bound to a 23 kDa polypeptide of lipid droplets that was not detected with the GTP gamma S binding assay. A 25 kDa component of milk lipid globules was a potent substrate for ADP-ribosylation by botulinum toxin C3, but cholera toxin was much less effective, suggesting that this component may belong to the rac class of G-proteins. The 21 kDa component was related immunologically to ADP ribosylation factor.

Milk lipid globule precursor release from endoplasmic reticulum reconstituted in a cell-free system.

Lipid droplet precursors of milk lipid globules are believed to be derived from elements of endoplasmic reticulum in milk-secreting mammary epithelial cells. Endoplasmic reticulum isolated from mammary gland was able to generate small droplets morphologically resembling microlipid droplet precursors of milk lipid globules. Droplet generation was time and temperature dependent and required a cytosol fraction of Mr greater than 10,000. Droplet generation was enhanced by, but did not require, addition of nucleoside triphosphates, fatty acids, coenzyme A, glycerol-3-phosphate, and dithiothreitol. Microlipid droplets generated in this cell-free system were enriched in triacylglycerols and resembled microlipid droplets formed within mammary epithelial cells in polar lipid and polypeptide composition. Endoplasmic reticulum immobilized onto nitrocellulose retained activity in generation of putative microlipid droplets, and this immobilization method provided a facile means for separation of the donor from the generated products.

Operon structure and nucleotide homology of the chlorocatechol oxidation genes of plasmids pJP4 and pAC27.

Alcaligenes eutrophus harboring plasmid pJP4 (strain JMP134) is capable of growing on both 2,4-dichlorophenoxyacetate (2,4-D) and 3-chlorobenzoate (3-Cba), while Pseudomonas putida carrying plasmid pAC27 (strain AC867) can utilize only 3-Cba as the sole carbon source. The tfdCDEF operon of the pJP4 plasmid and the clcABD operon of plasmid pAC27 each encode enzymes for the degradation of chlorocatechols (Clc), key intermediates in the catabolism of 2,4-D and 3-Cba. Similarities in the nucleotide (nt) sequences of genes tfdC and clcA, encoding pyrocatechases, were reported earlier [Ghosal and You, Mol. Gen. Genet. 211 (1988a) 113-120]. Genes tfdD and clcB, encoding Clc-specific cycloisomerases, have been completely sequenced. The tfdD gene (1107 bp) is slightly smaller than gene clcB (1113 bp). Comparison of the two cycloisomerase-encoding genes reveals that the nt sequences are 63% homologous with 62% homology in the deduced amino acid (aa) sequences of the polypeptides they encode. Genes tfdD and tfdE are contiguous in the tfdCDEF operon, whereas the corresponding genes, clcB and clcD, of the clcABD operon, are known to be separated by a long open reading frame of unknown function. The predicted N-terminal aa sequences of the two hydrolase-encoding genes, tfdE and clcD, also show homology. The structural and nt homologies between the two Clc operons, tfdCDEF and clcABD, suggest their relatedness.

Nucleotide sequence and expression of gene nahH of plasmid NAH7 and homology with gene xylE of TOL pWWO.

The enzyme catechol 2,3-dioxygenase (C23O) encoded by the nahH gene of plasmid NAH7 converts catechol to alpha-hydroxymuconic epsilon-semialdehyde in Pseudomonas putida. We have cloned this structural gene into vectors pUC18 and pKT240, determined the nucleotide sequence and deduced the amino acid sequence. In comparison to the gene xylE of the TOL plasmid pWWO which encodes a similar C230 enzyme [Nakai et al. J. Biol. Chem. (1983b), 2923-2928], the respective G + C contents were 55% and 57%, the nucleotide sequences 81% homologous, and the amino acid homology 85%. The flanking sequences of the two C23O-coding genes also show homology. Clones of the nahH gene in Escherichia coli overproduce the protein product at least ten fold and the gene product was identified by polyacrylamide gel electrophoresis.

IS2-61 and IS2-611 arise by illegitimate recombination from IS2-6.

A more stable derivative of IS2-6 has been isolated, which had lost 54 bp of the 108 bp long insert characteristic of IS2-6. This new allele of IS2, IS2-61, segregates the remaining 54 bp to yield allele IS2-611. DNA sequence analysis shows that the segregation products of IS2-6 arise by recA-independent, illegitimate recombination at 9 bp long direct sequence repetitions.

DNA sequence of the mini-insertion IS2--6 and its relation to the sequence of IS2.

In polar IS2 abolishes galactose operon expression. Operon activity is restored by a 108 base pair mini-insertion within IS2 called IS2--6. The DNA sequences of the gal operon-IS2 junction, the parental IS2 region undergoing sequence rearrangements and IS2--6 itself are reported. IS2--6 is composed of sequence intervals present in both strands of IS2.

Nucleotide homology and organization of chlorocatechol oxidation genes of plasmids pJP4 and pAC27.

The 2,4-dichlorophenoxyacetate (2,4-D) catabolic plasmid pJP4 of Alcaligenes eutrophus JMP134 contains two sets of nonidentical chlorocatechol oxidation gene sequences physically separated by a 7 kb DNA region. We determined the nucleotide sequence of the 1.6 kb HindIII fragment containing the known genes tfdC and tfdD (Don et al. 1985) which encode pyrocatechase and cycloisomerase, respectively. The 1.3 kb BglII-HindIII segment of recombinant plasmid pDC25 containing at least three chlorocatechol (clc) oxidation genes of the pAC27 plasmid in Pseudomonas putida AC867 (Ghosal et al. 1985a; Frantz and Chakrabarty 1986), was also sequenced. When the tfdC gene of the pJP4 plasmid was compared with gene clcA of plasmid pAC27, which encodes the chlorocatechol specific pyrocatechase (pyrocatechase II), the two genes showed 63% nucleotide sequence homology with 60% homology in their amino acid sequences. In both plasmid pJP4 and pAC27, the two genes encoding the pyrocatechase and the cycloisomerase showed a 4 bp overlap spanning the initiation codon of the cycloisomerase gene and the termination codon of the pyrocatechase gene. The sizes of the polypeptides encoded by the isofunctional genes tfdC and clcA are very similar and thus reflect their functional homology.

Sulfated macromolecules in early embryos of Limnaea.

An attempt has been made to partially characterize the substance responsible for the rhythmic incorporation of 35S (inorganic sulfate) in the course of the cell cycle in early Limnaea embryos. This trichloroacetic acid (TCA) insoluble substance is partly pronase sensitive, and dissociable into two fractions after treatment with NaCl. One of these remains TCA-insoluble while the other is TCA soluble but precipitable by cetyl pyridinium chloride. Thus, unlike in some other higher organisms, the major part of the inorganic sulfate is incorporated here into a fraction which is not a single mycopolysaccharide, but is more likely to be a protein-mucopolysaccharide complex, rather like the chondromucoprotein of chick-embryo cartilage.

Genetic and molecular analysis of a regulatory region of the herbicide 2,4-dichlorophenoxyacetate catabolic plasmid pJP4.

In Alcaligenes eutrophus JMP134, pJP4 carries the genes coding for 2,4-dichlorophenoxyacetate (2,4-D) and 3-chlorobenzoate (3-Cba) degradation plus mercury resistance. The plasmid genes specifying 2,4-D and 3-Cba catabolism are organized in three operons: tfdA, tfdB, and tfdCDEF. Regulation of these operons by two unlinked genes, tfdR and tfdS, has been proposed. Physical and DNA sequence analyses revealed that the tfdR and tfdS genes were identical and were located within a longer inverted repeat of 1592 bp. Similar stem-loop structures were observed among other 2,4-D plasmids. The tfdR gene is 888 bp long and capable of encoding a polypeptide of 32 kDa. The deduced amino acid sequence of tfdR indicates that it is a member of the LysR-type activators. Investigation of the regulation of the catabolic gene clusters through the construction of a pJP4 defined deletion mutant, pYG1010, which lacks a 4.2 kilobase Xbal fragment containing the inverted repeat region carrying the tfdR and tfdS regulatory genes, showed that Pseudomonas cepacia strains containing pYG1010 became 2,4-D negative, but 3-Cba positive. In vivo recombinants of pYG1010 and a cloned tfdS gene rescued the 2,4-D phenotype, indicating that TfdS is a positive regulator of tfdA expression, but not for tfdCDEF expression.

Multiple integration sites for the lactose transposon Tn 951 on plasmid RP 1 and establishment of a coordinate system for Tn 951.

Various molecules generated by transposition of the lactose transposon Tn 951 from plasmid pGC1 to plasmid RP1 were examined by DNA heteroduplex and restriction endonuclease analysis. Tn 951 was found to transpose to at least eight different sites on RP 1 in both possible orientations. A coordinate system for the lactose transposon Tn 951 is constructed.

Nucleotide sequence of the transposable DNA-element IS2.

The complete sequence of the transposable DNA element IS2 in gal OP-308:: IS2 (I) has been determined. This element is 1.327 bp long. The integrated element is flanked by a five base pair long sequence duplication. The termini of IS2 are not perfect inverted repeats, but a close approximation.

Gene duplication in haloaromatic degradative plasmids pJP4 and pJP2.

pJP2 and pJP4 are 2,4-dichlorophenoxyacetic acid catabolic plasmids, and they show DNA sequence homology. Most of the pJP2 molecules (80% or more) isolated from 2,4-dichlorophenoxyacetic acid grown cells of Alcaligenes eutrophus harbor a tandem duplication of a 25-kilobase (kb) segment encoding the catabolic functions. Unlike plasmid pJP4, pJP2 in A. eutrophus gives rise to a 3-chlorobenzoate phenotype without further genetic rearrangement. pJP4 under 3-chlorobenzoate selection contains an inverted duplication of 24.5 kb. Absence of selective pressure results in the prompt loss of one copy of the duplication in pJP4, but not of the tandem duplication in pJP2. In both pJP4 and pJP2, mutation of the duplicated copy, rather than gene dosage, is likely to be the basis of phenotypic change of catabolic functions. Experiments using the cloned DNA suggest that a tandem duplication is more stable than an inverted duplication.