bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural and optical properties of 1.3 μm wavelength InAsP/InP/InGaP strain-compensated multiple quantum well modulators grown by gas-source molecular beam epitaxy
Authors: Kuo H. C. , Thomas S. , Horton T. U. , Moser B. G. , Stillman G. E. , Lin C. H. , Chen H.
Published: July 2002

We present in this article a comparison of 1.3 μm InAs0.45P0.55/In0.81Ga0.19P strain-compensated multiple quantum well (SC-MQW) modulators with and without an InP insertion layer. Material quality was evaluated by cross-sectional transmission electron microscope analysis, x-ray rocking curves with dynamical simulations, and photoluminescence measurements. Devices were fabricated and device performance criteria such as contrast ratio (C.R.) and uniformity were also compared. It was found that higher C.R. and highly uniform InAsP/InGaP SC-MQW modulators can be achieved by inserting thin InP layers at the heterointerfaces between InAsP and InGaP.
Conformal ion implantation using pulsed plasma sources
Authors: Adler R. J. , Richter-Sand R. J. , Clark E. J. , Gregg C. W.
Published: July 2002

Operation of plasma implantation equipment at currents of several hundred amperes has been achieved with high density (1011–1012/cc) pulsed plasma sources. These currents have lead to inferred sheath dimensions in the 1–2 cm range. The equipment, results, and analysis are described here in detail.
Novel metallization technique for filling 100-nm-wide trenches and vias with very high aspect ratio
Authors: Monteiro Othon R.
Published: July 2002

We have developed a metallization method that successfully satisfies all the requirements for the next several generations of integrated circuits (ICs). This method was used to deposit fully conformal diffusion barriers of Ta and TaN as well as Cu seeding layers in 100-nm-wide trenches and vias with (depth:width) aspect ratios up to 8:1. The diffusion barrier and seeding layer are critical for the implementation of copper metallization schemes. This technique was also used to completely fill high aspect ratio trenches with copper. A fully ionized metal plasma deposition method is used in which the ion energy is varied and controlled as a function of time throughout the deposition process. No limitation of the process with respect to trench width or aspect ratio has been observed yet, suggesting the process can be used for ICs with sub-100 nm linewidths.
Control of seed layer for a low temperature formation of polycrystalline silicon with high crystallinity and a smooth surface
Authors: Murata Kazuya , Ito Masafumi , Hori Masaru , Goto Toshio
Published: July 2002

We have successfully improved the surface roughness and crystalline fraction of polycrystalline silicon (poly-Si) films formed on quartz substrates at 300 °C by controlling its initial growth. Seed layers were formed by eliminating charged species incident on the substrate during the initial growth in electron cyclotron resonance silane and hydrogen plasma-enhanced chemical vapor deposition and poly-Si films were subsequently formed on the seed layers with charged species. As a result, control of ion bombardment on the initial growth was found to be a key factor for forming poly-Si films with smooth surfaces and high crystallinity at low temperatures.
Photonic band-gap waveguide microcavities: Monorails and air bridges
Authors: Lim Kuo-Yi , Ripin D. J. , Petrich G. S. , Kolodziejski L. A. , Ippen E. P. , Mondol M. , Smith Henry I. , Villeneuve P. R. , Fan S. , Joannopoulos J. D.
Published: July 2002

Photonic band-gap monorail and air-bridge waveguide microcavities, operating at the wavelength regime of 1550 nm, are fabricated using GaAs-based compound semiconductors. The fabrication process involves gas-source molecular beam epitaxy, electron-beam lithography, reactive ion etching, and thermal wet oxidation of Al0.93Ga0.07As. The fabrication of the air-bridge microcavity, in particular, also entails the sacrificial wet etch of AlxOy to suspend the micromechanical structure. The monorail and air-bridge microcavities have been optically characterized and the transmission spectra exhibit resonances in the 1550 nm wavelength regime. Tunability of the resonant wavelength is demonstrated through changing the defect size in the one-dimensional photonic crystal. The quality factors (Q) of the microcavities are about 140 for the monorail and 230 for the air bridge, respectively.
Quantum dot-like behavior of GaInNAs in GaInNAs/GaAs quantum wells grown by gas-source molecular-beam epitaxy
Authors: Xin H. P. , Kavanagh K. L. , Zhu Z. Q. , Tu C. W.
Published: July 2002

A quantum dot-like behavior of GaInNAs in Ga0.7In0.3NxAs1−x/GaAs quantum wells (QWs) grown by gas-source molecular-beam epitaxy has been studied using high-resolution x-ray rocking curves (XRC), cross-sectional transmission electron microscopy (XTEM), and photoluminescence (PL) spectroscopy. XRC determines the average alloy composition and thickness of the QWs to be In 0.30 and N 0–0.030, and 6.2 nm, respectively. XTEM images show that the wells of both Ga0.7In0.3As/GaAs and Ga0.7In0.3N0.02As0.98/GaAs are undulated with lateral variations in strain, but the latter is much rougher. For Ga0.7In0.3As/GaAs QWs, rapid thermal annealing results in the splitting of a broad excitonic emission into two peaks presumably due to In composition fluctuation. The separation between these two peaks increases with increasing N concentration. Increasing excitation intensity results in a significant blueshift for the low-energy peak, while little blueshift for the high-energy peak. The earlier PL behavior can be interpreted by a simple model where the low-energy peak originates from In and N-rich regions in the wells acting as quantum dots. The high-energy peak is likely due to the excitons of a more two-dimensional QW.
Infrared and Raman studies of confined and interface optical phonons in short-period GaAs/AlAs superlattices with a grating coupler
Authors: Milekhin A. , Rösch M. , Batke E. , Zahn D. R. T. , Köhler K. , Ganser P. , Preobrazhenskii V. , Semyagin B.
Published: July 2002

The optical phonon modes in short-period GaAs/AlAs superlattices were investigated by means of Raman spectroscopy and far-infrared reflection spectroscopy using a grating coupler technique. Grating couplers provide a wave vector transfer parallel to the layers, enabling study of in-plane dispersion of the phonon modes in the superlattice structure. The mixing of confined and interface optical modes was observed as the in-plane phonon wave vector was varied. The experimental frequencies of the superlattice optical modes obtained from infrared and Raman spectra are in good agreement with calculations performed using a dielectric continuum model.
Tantalum and tantalum nitride films deposited by electron cyclotron resonance sputtering as barriers to copper diffusion
Authors: Ono Toshiro , Amazawa Takao , Nishimura Hiroshi , Matsuo Seitaro
Published: July 2002

Tantalum (Ta) and tantalum nitride (TaN) films deposited by electron cyclotron resonance (ECR) sputtering have been investigated as barriers to Cu diffusion in very large scale integrated interconnections. An ECR plasma source coupled with divided microwaves was used to deposit Ta and TaN films. The addition of N2 to Ar enabled the ECR TaN film to be easily deposited by using a Ta metal target at low temperature without external substrate heating. To investigate the barrier characteristics, sample with Cu/ECR–Ta/SiO2 and Cu/ECR–TaN/SiO2 structures were heat treated in Ar at 550 and 650 °C, respectively, and analyzed by secondary ion mass spectroscopy, which determined the Cu diffusion density. The normalized signal intensity ratio of Cu to Si at the interface between the barrier metal and SiO2 was 0.035 for the ECR TaN film and 0.087 for the ECR Ta film, but 0.26 for the rf-sputtered TaN film. ECR sputter deposited films had excellent characteristics for use as barriers to Cu diffusion.
Low energy electron-beam proximity projection lithography: Discovery of a missing link
Authors: Utsumi Takao
Published: July 2002

Low energy electron(e)-beam proximity projection lithography is proposed for integrated circuit lithography for minimum feature sizes ⩽0.1 μm. This new e-beam lithography is similar to optical projection lithography except that photons are replaced by low energy electrons of 2 kV. The low e-beam energy permits the use of single crystal 0.5 μm thick silicon membrane masks without an absorbing metal layer of high atomic number. This membrane mask is thick enough for good heat conduction and thin enough for feature sizes ⩽0.1 μm. The mask distortion caused by the fabrication can be corrected by a fine-tuning deflector. Therefore the mask is allowed to have a residual distortion of more than 100 nm. The proposed system does not suffer from the space charge effect in the electron optics column nor the proximity effect with respect to both the wafer and mask writings, and it is fundamentally a low power lithography which needs no special cooling system. The analysis shows that the e-beam column can be made entirely of electrostatic components to achieve sufficient resolution. For an appropriate resist process for this low energy e-beam we propose a bilayer process which consists of a chemically amplified thin deep ultraviolet photoresist and a thick antireflective layer. We estimated a throughput of 30 12 in. wafers per hour and a resolution of less than 50 nm.
Stamp technology for fabrication of field emitter from organic material
Authors: Baba Akiyoshi , Hizukuri Masafumi , Iwamoto Masakazu , Asano Tanemasa
Published: July 2002

We propose and demonstrate a novel processing technique for fabrication of field electron emitter arrays. A stamp technology was used, in which the emitter tips are prepared by transferring the shape of a mold to an organic material through pressing the mold against the organic material. The transformed organic material is then modified by ion irradiation to produce a carbon based emitter material. The starting material tested was a polyimide film spin coated and cured on a Si substrate. The mold was prepared using anisotropic etching of Si(100). It has been found that the mold shape can be completely transferred to the polyimide by applying a pressure of 1 GPa at 250 °C. A field emission current up to the order of μA was obtained from a wedge-shaped emitter array made using this new technology with modification using Ar ion irradiation at an energy of 100 keV, a dose of 3×1016 cm−2.
Effects of substrate bias on the structural and field emission properties of diamond films
Authors: Shim Jae Yeob , Chi Eung Joon , Baik Hong Koo , Song Kie Moon , Lee Se-Jong
Published: July 2002

The effect of electron bombardment on modification of the structural property and the surface morphology of diamond films, and consequently the field emission properties have been investigated by introducing positive substrate bias. When increasing the bias voltage, the structural properties of diamond films are significantly deteriorated together with the increase of nondiamond carbon component and the surface morphologies of the films lost their unique facet shape. The reason for the increase of nondiamond carbon content is described in terms of both the increase of substrate temperature and the excessive generation of CHn radicals. The field emission properties of diamond films were substantially improved with increasing bias voltage. Enhancement of field emission property for the films prepared by increasing bias voltage and/or methane concentration has been discussed by correlating between the substrate–diamond interface property and the slope of the Fowler–Nordheim plot.
Promoter Recognition As Measured by Binding of Polymerase to Nontemplate Strand Oligonucleotide
Authors: Marr M. T. , Roberts J. W.
Published: July 2002

In transcription initiation, the DNA strands must be separated to expose the template to RNA polymerase. As the closed initiation complex is converted to an open one, specific protein-DNA interactions involving bases of the nontemplate strand form and stabilize the promoter complex in the region of unwinding. Specific interaction between RNA polymerase and the promoter in
Escherichia coli
was detected and quantified as the binding affinity of nontemplate oligonucleotide sequences. The RNA polymerase subunit sigma factor 70 contacted the bases of the nontemplate DNA strand through its conserved region 2; a mutation that affected promoter function altered the binding affinity of the oligonucleotide to the enzyme.
Effects of Random Perturbations in Plastic Optical Fibers
Authors: Garito A. F. , Wang J. , Gao R.
Published: July 2002

REVIEW

The most important feature of an optical fiber waveguide is its bandwidth, which defines its information-carrying capacity. A major limitation on the bandwidth of multimode glass and plastic optical fibers is modal dispersion, in which different optical modes propagate at different velocities and the dispersion grows linearly with length. However, in plastic optical fibers, experimental and theoretical results indicate that the modes are not independent but are highly coupled, which leads to a characteristic square-root length dependence and an unanticipated large enhancement of the bandwidth to gigahertz levels. The ever increasing demands for low-cost, high-bandwidth communications media for voice, video, and data transmission in short- and medium-distance applications are generating a new assessment of multimode optical fibers to serve as high-speed fiber links.
Activities and Postantibiotic Effects of Gemifloxacin Compared to Those of 11 Other Agents against
Haemophilus influenzae
and
Moraxella catarrhalis
Authors: Davies Todd A. , Kelly Linda M. , Hoellman Dianne B. , Ednie Lois M. , Clark Catherine L. , Bajaksouzian Saralee , Jacobs Michael R. , Appelbaum Peter C.
Published: July 2002

The activity of gemifloxacin against
Haemophilus influenzae
and
Moraxella catarrhalis
was compared to those of 11 other agents. All quinolones were very active (MICs, ≤0.125 μg/ml) against 248 quinolone-susceptible
H. influenzae
isolates (40.7% of which were β-lactamase positive); cefixime (MICs, ≤0.125 μg/ml) and amoxicillin-clavulanate (MICs ≤4.0 μg/ml) were active, followed by cefuroxime (MICs, ≤16.0 μg/ml); azithromycin MICs were ≤4.0 μg/ml. For nine
H. influenzae
isolates with reduced quinolone susceptibilities, the MICs at which 50% of isolates are inhibited (MIC
50
s) were 0.25 μg/ml for gemifloxacin and 1.0 μg/ml for the other quinolones tested. All strains had mutations in GyrA (Ser84, Asp88); most also had mutations in ParC (Asp83, Ser84, Glu88) and ParE (Asp420, Ser458), and only one had a mutation in GyrB (Gln468). All quinolones tested were equally active (MICs, ≤0.06 μg/ml) against 50
M. catarrhalis
strains; amoxicillin-clavulanate, cefixime, cefuroxime, and azithromycin were very active. Against 10
H. influenzae
strains gemifloxacin, levofloxacin, sparfloxacin, and trovafloxacin at 2× the MIC and ciprofloxacin at 4× the MIC were uniformly bactericidal after 24 h, and against 9 of 10 strains grepafloxacin at 2× the MIC was bactericidal after 24 h. After 24 h bactericidal activity was seen with amoxicillin-clavulanate at 2× the MIC for all strains, cefixime at 2× the MIC for 9 of 10 strains, cefuroxime at 4× the MIC for all strains, and azithromycin at 2× the MIC for all strains. All quinolones except grepafloxacin (which was bactericidal against four of five strains) and all ß-lactams at 2× to 4× the MIC were bactericidal against five
M. catarrhalis
strains after 24 h; azithromycin at the MIC was bactericidal against all strains after 24 h. The postantibiotic effects (PAEs) against four quinolone-susceptible
H. influenzae
strains were as follows: gemifloxacin, 0.3 to 2.3 h; ciprofloxacin, 1.3 to 4.2 h; levofloxacin, 2.8 to 6.2 h; sparfloxacin, 0.6 to 3.0 h; grepafloxacin, 0 to 2.1 h; trovafloxacin, 0.8 to 2.8 h. At 10× the MIC, no quinolone PAEs were found against the strain for which quinolone MICs were increased. Azithromycin PAEs were 3.7 to 7.3 h.
Antagonism between Aminoglycosides and β-Lactams in a Methicillin-Resistant
Staphylococcus aureus
Isolate Involves Induction of an Aminoglycoside-Modifying Enzyme
Authors: Ida Takashi , Okamoto Ryoichi , Nonoyama Masato , Irinoda Kazuhiko , Kurazono Mizuyo , Inoue Matsuhisa
Published: July 2002

We encountered three clinical isolates of methicillin-resistant
Staphylococcus aureus
which were susceptible to netilmicin and arbekacin in the absence of β-lactam antibiotics but which were resistant to them in the presence of β-lactam antibiotics. One of these strains, KU5801, was used to further investigate the antagonism between aminoglycosides and β-lactam antibiotics. β-Lactam antibiotics induced bacterial synthesis of aminoglycoside-6′-
N
-acetyltransferase and 2"-
O
-phosphotransferase [AAC(6′)-APH(2")] in association with decreased antimicrobial activities of aminoglycosides. A 14.4-kb
Eco
RI fragment that included the genes that control for β-lactam-inducible aminoglycoside resistance was cloned from a 31-kb conjugative plasmid present in KU5801. Restriction fragment mapping and PCR analysis suggested that a Tn
4001
-like element containing a gene encoding AAC(6′)-APH(2") was located downstream from a truncated
blaZ
gene. The DNA sequence between
blaR1
and a Tn
4001
-like element was determined. The Tn
4001
-IS
257
hybrid structure was cointegrated into the
blaZ
gene, and the typical sequences for the termination of transcription were not found between these regions. We deduced that antagonism of aminoglycosides by β-lactam antibiotics in isolate KU5801 involved transcription of the
aac(6′)-Ie-aph(2")-Ia
gene under the influence of the system regulating penicillinase production.
Molecular and Physiological Responses of Two Classes of Marine Chromophytic Phytoplankton (Diatoms and Prymnesiophytes) during the Development of Nutrient-Stimulated Blooms
Authors: Wyman Michael , Davies John T. , Crawford David W. , Purdie Duncan A.
Published: July 2002

Generic taxon-specific DNA probes that target an internal region of the gene (
rbcL
) encoding the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) were developed for two groups of marine phytoplankton (diatoms and prymnesiophytes). The specificity and utility of the probes were evaluated in the laboratory and also during a 1-month mesocosm experiment in which we investigated the temporal variability in RubisCO gene expression and primary production in response to inorganic nutrient enrichment. We found that the onset of successive bloom events dominated by each of the two classes of chromophyte algae was associated with marked taxon-specific increases in
rbcL
transcription rates. These observations suggest that measurements of RubisCO gene expression can provide an early indicator of the development of phytoplankton blooms and may also be useful in predicting which taxa are likely to dominate a bloom.
Consumption of Tropospheric Levels of Methyl Bromide by C
1
Compound-Utilizing Bacteria and Comparison to Saturation Kinetics
Authors: Goodwin Kelly D. , Varner Ruth K. , Crill Patrick M. , Oremland Ronald S.
Published: July 2002

Pure cultures of methylotrophs and methanotrophs are known to oxidize methyl bromide (MeBr); however, their ability to oxidize tropospheric concentrations (parts per trillion by volume [pptv]) has not been tested. Methylotrophs and methanotrophs were able to consume MeBr provided at levels that mimicked the tropospheric mixing ratio of MeBr (12 pptv) at equilibrium with surface waters (≈2 pM). Kinetic investigations using picomolar concentrations of MeBr in a continuously stirred tank reactor (CSTR) were performed using strain IMB-1 and
Leisingeria methylohalidivorans
strain MB2
T
— terrestrial and marine methylotrophs capable of halorespiration. First-order uptake of MeBr with no indication of threshold was observed for both strains. Strain MB2
T
displayed saturation kinetics in batch experiments using micromolar MeBr concentrations, with an apparent
K

s

of 2.4 μM MeBr and a
V
max
of 1.6 nmol h
−1
(10
6
cells)
−1
. Apparent first-order degradation rate constants measured with the CSTR were consistent with kinetic parameters determined in batch experiments, which used 35- to 1 × 10
7
-fold-higher MeBr concentrations.
Ruegeria algicola
(a phylogenetic relative of strain MB2
T
), the common heterotrophs
Escherichia coli
and
Bacillus pumilus
, and a toluene oxidizer,
Pseudomonas mendocina
KR1, were also tested. These bacteria showed no significant consumption of 12 pptv MeBr; thus, the ability to consume ambient mixing ratios of MeBr was limited to C
1
compound-oxidizing bacteria in this study. Aerobic C
1
bacteria may provide model organisms for the biological oxidation of tropospheric MeBr in soils and waters.
Attenuated Nontoxinogenic and Nonencapsulated Recombinant
Bacillus anthracis
Spore Vaccines Protect against Anthrax
Authors: Cohen S. , Mendelson I. , Altboum Z. , Kobiler D. , Elhanany E. , Bino T. , Leitner M. , Inbar I. , Rosenberg H. , Gozes Y. , Barak R. , Fisher M. , Kronman C. , Velan B. , Shafferman A.
Published: July 2002

Several highly attenuated spore-forming nontoxinogenic and nonencapsulated
Bacillus anthracis
vaccines differing in levels of expression of recombinant protective antigen (rPA) were constructed. Biochemical analyses (including electrospray mass spectroscopy and N terminus amino acid sequencing) as well as biological and immunological tests demonstrated that the rPA retains the characteristics of native PA. A single immunization of guinea pigs with 5 × 10
7
spores of one of these recombinant strains, MASC-10, expressing high levels of rPA (≥100 μg/ml) from a constitutive heterologous promoter induced high titers of neutralizing anti-PA antibodies. This immune response was long lasting (at least 12 months) and provided protection against a lethal challenge of virulent (Vollum) anthrax spores. The recombinant
B. anthracis
spore vaccine appears to be more efficacious than the vegetative cell vaccine. Furthermore, while results clearly suggest a direct correlation between the level of expression of PA and the potency of the vaccine, they also suggest that some
B. anthracis
spore-associated antigen(s) may contribute in a significant manner to protective immunity.
Genetic Polymorphisms of Group B StreptococcusscpBAlter Functional Activity of a Cell-Associated Peptidase That Inactivates C5a
Authors: Bohnsack John F. , Takahashi Shinji , Hammitt Laura , Miller Dylan V. , Aly Adrienne A. , Adderson Elisabeth E.
Published: July 2002

Many group BStreptococcus agalactiaestrains and other pathogenic streptococci express a cell-associated peptidase that inactivates C5a (C5a-ase), the major neutrophil chemoattractant produced by activation of the complement cascade. Type III group B streptococci (GBS) can be classified genotypically into three restriction digest pattern types. Functional C5a-ase activity of GBS correlates with this genetic typing; therefore, we sought to identify a genetic basis for this phenomenon. Southern hybridization confirms that all type III GBS containscpB, the gene encoding GBS C5a-ase. GBS strains with high C5a-ase functional activity and those with no or very low activity both express immunoreactive C5a-ase. ThescpBsequence of strain I30, which has high C5a-ase activity, is 98.2% homologous to the previously reported serotype II GBSscpBsequence. ThescpBsequences of strains I25 and GW, which have low or no C5a-ase activity, are identical. The predicted I25 and GW C5a-ase proteins share a four-amino-acid deletion affecting the protease histidine active-site consensus motif. Recombinant I30 C5a-ase has good functional activity, whereas recombinant I25 C5a-ase has low activity. These data demonstrate that functional C5a-ase differences between type III GBS strains are attributable to a genetic polymorphism ofscpB. The ubiquitous expression of C5a-ase, irrespective of functional activity, suggests that C5a-ase may have a second, as yet unidentified, function.
Differential Vβ T-Cell Receptor Usage during Chronic Experimental Schistosomiasis Corresponds with Distinct Pathological Presentations
Authors: Secor W. Evan , Freeman George L.
Published: July 2002

CBA/J male mice with chronicSchistosoma mansoniinfections display either moderate splenomegaly syndrome (MSS) or hypersplenomegaly syndrome (HSS). As MSS and HSS mice differ in several immunologic characteristics, we investigated T-cell receptor Vβ usage. The groups had significantly different expression of several Vβs, suggesting a relationship between the T-cell repertoire and schistosomiasis pathology.
Mice Lacking T and B Lymphocytes Develop Transient Colitis and Crypt Hyperplasia yet Suffer Impaired Bacterial Clearance during
Citrobacter rodentium
Infection
Authors: Vallance Bruce A. , Deng Wanyin , Knodler Leigh A. , Finlay B. Brett
Published: July 2002

The bacterial pathogen
Citrobacter rodentium
belongs to a family of gastrointestinal pathogens that includes enteropathogenic and enterohemorrhagic
Escherichia coli
and is the causative agent of transmissible colonic hyperplasia in mice. The molecular mechanisms used by these pathogens to colonize host epithelial surfaces and form attaching and effacing (A/E) lesions have undergone intense study. In contrast, little is known about the host's immune response to these infections and its importance in tissue pathology and bacterial clearance. To address these issues, wild-type mice and mice lacking T and B lymphocytes (RAG1 knockout [KO]) were infected with
C. rodentium.
By day 10 postinfection (p.i.), both wild-type and RAG1 KO mice developed colitis and crypt hyperplasia, and these responses became more exaggerated in wild-type mice over the next 2 weeks, as they cleared the infection. By day 24 p.i., bacterial clearance was complete, and the colitis had subsided; however, crypt heights remained increased. In contrast, inflammatory and crypt hyperplastic responses in the RAG1 KO mice were transient, subsiding after 2 weeks. By day 24 p.i., RAG1 KO mice showed no signs of bacterial clearance and infection was often fatal. Surprisingly, despite remaining heavily infected, tissues from RAG1 KO mice surviving the acute colitis showed few signs of disease. These results thus emphasize the important contribution of the host immune response during infection by A/E bacterial pathogens. While T and/or B lymphocytes are essential for host defense against
C. rodentium
, they also mediate much of the tissue pathology and disease symptoms that occur during infection.
Functional Analysis of 14 Genes That Constitute the Purine Catabolic Pathway in
Bacillus subtilis
and Evidence for a Novel Regulon Controlled by the PucR Transcription Activator
Authors: Schultz Anna C. , Nygaard Per , Saxild Hans H.
Published: July 2002

The soil bacterium
Bacillus subtilis
has developed a highly controlled system for the utilization of a diverse array of low-molecular-weight compounds as a nitrogen source when the preferred nitrogen sources, e.g., glutamate plus ammonia, are exhausted. We have identified such a system for the utilization of purines as nitrogen source in
B. subtilis
. Based on growth studies of strains with knockout mutations in genes, complemented with enzyme analysis, we could ascribe functions to 14 genes encoding enzymes or proteins of the purine degradation pathway. A functional xanthine dehydrogenase requires expression of five genes (
pucA, pucB, pucC, pucD
, and
pucE
). Uricase activity is encoded by the
pucL
and
pucM
genes, and a uric acid transport system is encoded by
pucJ
and
pucK
. Allantoinase is encoded by the
pucH
gene, and allantoin permease is encoded by the
pucI
gene. Allantoate amidohydrolase is encoded by
pucF
. In a
pucR
mutant, the level of expression was low for all genes tested, indicating that PucR is a positive regulator of
puc
gene expression. All 14 genes except
pucI
are located in a gene cluster at 284 to 285° on the chromosome and are contained in six transcription units, which are expressed when cells are grown with glutamate as the nitrogen source (limiting conditions), but not when grown on glutamate plus ammonia (excess conditions). Our data suggest that the 14 genes and the
gde
gene, encoding guanine deaminase, constitute a regulon controlled by the
pucR
gene product. Allantoic acid, allantoin, and uric acid were all found to function as effector molecules for PucR-dependent regulation of
puc
gene expression. When cells were grown in the presence of glutamate plus allantoin, a 3- to 10-fold increase in expression was seen for most of the genes. However, expression of the
pucABCDE
unit was decreased 16-fold, while expression of
pucR
was decreased 4-fold in the presence of allantoin. We have identified genes of the purine degradation pathway in
B. subtilis
and showed that their expression is subject to both general nitrogen catabolite control and pathway-specific control.
Lambda Excision Revisited: Testing a Model for Synapsis of Prophage Ends
Authors: Pato Martin L.
Published: July 2002

Excision of lambda prophage was reexamined to test a model for prophage end synapsis. The model proposes that, during in situ prophage replication, following induction, the diverging replication forks are held together. Consequently, prophage DNA is spooled through the replication machinery, drawing the prophage ends together and facilitating synapsis. The model predicts that excision will be slowed if in situ lambda replication is inhibited, and the predicted low rate of excision of a nonreplicating prophage was observed after thermoinduction. However, excision was rapid if additional Int protein was supplied or if the temperature was reduced after induction, showing that (i) Int is partially thermosensitive for excision at 42°C and (ii) in situ replication is not required for rapid excision, a finding that is inconsistent with the model.
Functional Genomic, Biochemical, and Genetic Characterization of the
Salmonella pduO
Gene, an ATP:Cob(I)alamin Adenosyltransferase Gene
Authors: Johnson Celeste L. V. , Pechonick Edith , Park Sanghee D. , Havemann Gregory D. , Leal Nicole A. , Bobik Thomas A.
Published: July 2002

Salmonella enterica
degrades 1,2-propanediol by a pathway dependent on coenzyme B
12
(adenosylcobalamin [AdoCb1]). Previous studies showed that 1,2-propanediol utilization (
pdu
) genes include those for the conversion of inactive cobalamins, such as vitamin B
12
, to AdoCbl. However, the specific genes involved were not identified. Here we show that the
pduO
gene encodes a protein with ATP:cob(I)alamin adenosyltransferase activity. The main role of this protein is apparently the conversion of inactive cobalamins to AdoCbl for 1,2-propanediol degradation. Genetic tests showed that the function of the
pduO
gene was partially replaced by the
cobA
gene (a known ATP:corrinoid adenosyltransferase) but that optimal growth of
S. enterica
on 1,2-propanediol required a functional
pduO
gene. Growth studies showed that
cobA pduO
double mutants were unable to grow on 1,2-propanediol minimal medium supplemented with vitamin B
12
but were capable of growth on similar medium supplemented with AdoCbl. The
pduO
gene was cloned into a T7 expression vector. The PduO protein was overexpressed, partially purified, and, using an improved assay procedure, shown to have cob(I)alamin adenosyltransferase activity. Analysis of the genomic context of genes encoding PduO and related proteins indicated that particular adenosyltransferases tend to be specialized for particular AdoCbl-dependent enzymes or for the de novo synthesis of AdoCbl. Such analyses also indicated that PduO is a bifunctional enzyme. The possibility that genes of unknown function proximal to adenosyltransferase homologues represent previously unidentified AdoCbl-dependent enzymes is discussed.
Functional Role for a 2-Oxo Acid Dehydrogenase in the Halophilic Archaeon
Haloferax volcanii
Authors: Wanner Christian , Soppa Jörg
Published: July 2002

The archaeon
Haloferax volcanii
was previously shown to contain and transcribe the genes for a 2-oxo acid dehydrogenase (OADH) complex, but their presence remained a mystery because no enzymatic activity with any of the known OADH substrates could be found, and an inactivation of one of the genes did not lead to any phenotype. Here we report the identification of an additional
oadh
gene cluster in the genome of
H. volcanii
. In contrast to previously known
oadh
loci, it contains three genes,
oadh2A1
,
oadh2A2
, and
oadh2ld
, with coding capacity for the E1α and E1β subunits and an unattached lipoyl domain, but it is devoid of the genes for a complete E2 and an E3. The genes were isolated by complementation of a nitrate respiration-deficient mutant of
H. volcanii
and therefore were shown to be functional in vivo. Phylogenetic analyses revealed that the deduced E1α and E1β subunits of OADH2 group with bacterial acetoin dehydrogenases but not with the OADH1 subunits, and thus,
H. volcanii
has obtained the two gene groups independently. Comparison of the wild type and the mutant allowed us to exclude a function of OADH2 in the aerobic or anaerobic degradation of acetoin or glucose. Instead, it could be shown that OADH2 is important during nitrate-respirative growth on Casamino Acids. Many physiological and biochemical experiments failed to indicate that OADH2 uses any of the previously known OADH substrates. Growth potentials of the mutant were markedly different in media with a single carbon source versus media with mixed carbon sources.
Extensive Variation in the O-Antigen Gene Cluster within One
Salmonella enterica
Serogroup Reveals an Unexpected Complex History
Authors: Wang Lei , Andrianopoulos Kanella , Liu Dan , Popoff Michel Y. , Reeves Peter R.
Published: July 2002

The 46 serogroups of
Salmonella enterica
have different O-antigens, and each is thought to have a specific form of the O-antigen cluster. Comparison of the 145 serovars of serogroup B revealed much more intraserogroup genetic diversity than expected. The O27 factor, due to an α 1-6 linkage between O units in place of the more common α 1-2 linkage and previously thought to be due to a converting bacteriophage, is now shown to be due to a
wzy

α
(1-6)

gene located within the major gene cluster. Surprisingly a remnant of this gene in all O27
−
serovars shows that the ancestor was O27
+
. There are six distinct gene cluster forms, five apparently derived by a series of deletions and one by an insertion from an ancestral O27
+
form present in 57 serovars. The history of the gene cluster and movement between subspecies I and II can be traced. Two of the derivative forms still have a functional
wzy
α(1-6)
gene, while in three it has been inactivated by deletion or insertion. Two of the forms lacking a functional
wzy
α(1-6)
gene have the
wzy
α(1-2)
gene first described for strain LT2 as
rfc
, whereas for the third the
wzy
gene has not been located.
Identification of the cAD1 Sex Pheromone Precursor in
Enterococcus faecalis
Authors: An Florence Y. , Clewell Don B.
Published: July 2002

The
Enterococcus faecalis
virulence plasmid pAD1 encodes a mating response induced by exposure to an octapeptide sex pheromone, cAD1, secreted by plasmid-free enterococci. The determinant for the pheromone in
E. faecalis
FA2-2, designated
cad
, was found to encode a 309-amino-acid lipoprotein precursor with the last 8 residues of its 22-amino acid signal sequence representing the cAD1 moiety. The lipoprotein moiety contained two 77-amino-acid repeats (70% identity) separated by 45 residues. The nonisogenic
E. faecalis
strain V583 determinant encodes a homologous precursor protein, but it differs at two amino acid positions, both of which are located within the pheromone peptide moiety (positions 2 and 8). Construction of a variant of strain FA2-2 containing the differences present in V583 resulted in cells that did not produce detectable cAD1. The mutant appeared normal under laboratory growth conditions, and while significantly reduced in recipient potential, when carrying pAD1 it exhibited a normal mating response. A mutant of FA2-2 with a truncated lipoprotein moiety appeared normal with respect to recipient potential and, when carrying plasmid DNA, donor potential. A gene encoding a protein designated Eep, believed to be a zinc metalloprotease, had been previously identified as required for pheromone biosynthesis and was believed to be involved in the processing of a pheromone precursor. Our new observation that the pAD1-encoded inhibitor peptide, iAD1, whose precursor is itself a signal sequence, is also dependent on Eep is consistent with the likelihood that such processing occurs at the amino terminus of the cAD1 moiety.
Comparison of Pulsed-Field Gel Electrophoresis and Amplified Fragment Length Polymorphism Techniques for Investigating Outbreaks of Enteritis Due to Campylobacters
Authors: Champion Olivia L. , Best Emma L. , Frost Jennifer A.
Published: July 2002

Campylobacters are the most commonly reported cause of acute bacterial enteritis in the United Kingdom and United States, with poultry, milk, and water implicated as sources or vehicles of infection. The majority of campylobacter infections are sporadic, although outbreaks may occur, and these provide an opportunity to evaluate genotypic fingerprinting techniques. In this study, pulsed-field gel electrophoresis (PFGE) was compared with single-enzyme-amplified fragment length polymorphism (SAFLP). The results for the three separate episodes indicated that SAFLP and PFGE both clustered the strains from the first incident as 100% homologous. The strains from the second and third incidents clustered as distinct from both the first incident and from each other. PFGE is well recognized as a discriminatory fingerprinting technique for campylobacters; however, SAFLP has proven to be equally discriminatory, but far less labor intensive and with the added advantages of less “hands-on” time and inexpensive equipment, it is an excellent alternative to PFGE for investigation of outbreaks.
Alpha/Beta Interferon Promotes Transcription and Inhibits Replication of Borna Disease Virus in Persistently Infected Cells
Authors: Staeheli Peter , Sentandreu Maria , Pagenstecher Axel , Hausmann Jürgen
Published: July 2002

Borna disease virus(BDV) is a noncytolytic RNA virus that can replicate in the central nervous system (CNS) of mice. This study shows that BDV multiplication was efficiently blocked in transgenic mice that express mouse alpha-1 interferon (IFN-α1) in astrocytes. To investigate whether endogenous virus-induced IFN might similarly restrict BDV, we usedIFNAR0/0mice, which lack a functional alpha/beta IFN (IFN-α/β) receptor. As would be expected if virus-induced IFN were important to control BDV infection, we found that cultured embryo cells ofIFNAR0/0mice supported viral multiplication, whereas cells from wild-type mice did not. Unexpectedly, however, BDV spread through the CNSs ofIFNAR0/0and wild-type mice with similar kinetics, suggesting that activation of endogenous IFN-α/β genes in BDV-infected brains was too weak or occurred too late to be effective. Surprisingly, Northern blot analysis showed that the levels of the most abundant viral mRNAs in the brains of persistently infectedIFNAR0/0mice were about 20-fold lower than those in wild-type mice. In contrast, genomic viral RNA was produced in about a 10-fold excess in the brains ofIFNAR0/0mice. Human IFN-α2 similarly enhanced transcription and simultaneously repressed replication of the BDV genome in persistently infected Vero cells. Thus, in persistently infected neurons and cultured cells, IFN-α/β appears to freeze the BDV polymerase in the transcriptional mode, resulting in enhanced viral mRNA synthesis and suppressing viral genome replication.
Differentiation-Dependent Chromatin Rearrangement Coincides with Activation of Human Papillomavirus Type 31 Late Gene Expression
Authors: Peña Loren del Mar , Laimins Laimonis A.
Published: July 2002

The life cycle of human papillomaviruses (HPVs) is tightly linked to the differentiation status of the host cell. While early genes are expressed during the initial stages of viral infection, late gene expression occurs in the suprabasal layers of the cervical epithelium. Late genes encode E1^E4, a cytosolic protein, and capsid proteins L1 and L2. We have mapped over 30 initiation sites for late transcripts and show that the transcripts initiate in a 200-nucleotide region within the E7 open reading frame. The mechanisms regulating the activation of late gene expression, however, are not yet understood. DNase I hypersensitivity analysis of HPV-31 chromatin in cell lines that maintain viral genomes extrachromosomally indicates that a major shift in nuclease digestion occurs upon differentiation. In undifferentiated cells, hypersensitive regions exist in the upstream regulatory region proximal to the E6 open reading frame. Upon differentiation, a region between nucleotides 659 and 811 in the E7 open reading frame becomes accessible to DNase I. These results indicate that the late transcript initiation region becomes accessible to transcription factor binding upon differentiation. Several complexes mediate chromatin rearrangement, and we tested whether histone acetylation was sufficient for late transcript activation. Treatment with the histone deacetylase inhibitor trichostatin A was found to be insufficient to activate late gene expression in undifferentiated cells. However, it did activate expression of early transcripts. These results suggest that chromatin remodeling around the late promoter occurs upon epithelial differentiation and that mechanisms in addition to histone deacetylation contribute to activation of late gene expression.
Selection for Loss of p53 Function in T-Cell Lymphomagenesis Is Alleviated by Moloney Murine Leukemia Virus Infection in
myc
Transgenic Mice
Authors: Baxter Euan W. , Blyth Karen , Cameron Ewan R. , Neil James C.
Published: July 2002

Thymic lymphomas induced by Moloney murine leukemia virus (MMLV) have provided many examples of oncogene activation, but the role of tumor suppressor pathways in these tumors is less clear. These tumors display little evidence of loss of heterozygosity, and MMLV is only weakly synergistic with the
Trp53
null genotype, suggesting that viral lymphomagenesis involves mechanisms which do not require mutational loss of
Trp53
function. To explore this relationship in greater depth, we infected CD2-
myc
transgenic mice with MMLV and examined the role of
Trp53
in the genesis of these tumors. Most (19 of 27) of the tumors from MMLV-infected, CD2-
myc Trp53
+/−
mice retained the wild-type
Trp53
allele in vivo while tumors of uninfected CD2-
myc
Trp53
+/−
mice invariably showed allele loss from a significant fraction of primary tumor cells. The functional integrity of the
Trp53
gene in these tumors was indicated by ongoing allele loss or selection for mutational stabilization during in vitro propagation and by the radiosensitivity of selected
Trp53
+/−
tumor cell lines. An inverse correlation was noted between retention of the wild-type
Trp53
allele and expression of p19
ARF
, providing further evidence of negative-feedback control of the latter by p53. However, expression of p19
ARF
does not appear to be counterselected in the absence of p53, and its integrity in
Trp53
+/−
tumors was indicated by its transcriptional upregulation on
Trp53
wild-type allele loss in vitro in selected tumor cell lines. The role of MMLV was investigated further by analysis of proviral insertion sites in tumors of CD2-
myc
transgenic mice sorted for
Trp53
genotype. A proportion of tumors showed insertions at
Runx2
, an oncogene which has been shown to collaborate independently with CD2-
myc
and with the
Trp53
null genotype, and at a novel common integration site (
ptl-1
) on chromosome 8. Genotypic analysis of the panel of tumors suggested that neither of these integrations is functionally redundant with loss of p53, but it appears that the combination of the MMLV oncogenic program with the CD2-
myc
oncogene relegates p53 loss to a late step in tumor progression or in vitro culture. While the means by which these tumors preempt the p53 tumor suppressor response remains to be established, this study provides further evidence that irreversible inactivation of this pathway is not a prerequisite for tumor development in vivo.
Capillary Gas-Chromatographic Retention Behavior and Physico-Chemical Properties of Underivatized Equine Estrogens
Authors: Novakovic Jasmina , Tvrzická Eva , Všetečka Václav , Pouzar Vladimír , Feltl Ladislav
Published: July 2002

The capillary gas-chromatographic retention behavior of six underivatized equine estrogens (estrone, equilin, equilenin and their corresponding 17α-diols) on a mixed stationary phase (non-polar CP-Sil 5 CB and slightly polar CP-Sil 19 CB capillary columns coupled in series) was characterized by their Kovats retention indices and steroid numbers. In order to find whether a correlation exists between the chromatographic retention and some physico-chemical properties of the compounds, their octanol-water partition coefficients and dipole moments were measured. No straightforward correlation was observed between the physico-chemical properties and the retention behavior, which suggests that the GC separation of equine estrogens is governed by some specific electron donor-electron acceptor interactions of a chemical nature.
Synthesis and Kinetics of Isomerization of mer and fac[CoCO3(en)(Amino Acidato)] Isomers
Authors: Tyle Pavel , Jursík František
Published: July 2002

Reaction of [CoXY(en)(AB)]n+ (AB = glycinato or (S)-α-alaninato anion, X = H2O, Y = Cl; AB = (S)-valinato anion, X = Y = Cl) with HCO-3) or Ag(2CO)3 gives mixture of mer- and fac-[CoCO(3(en)(AB)] isomers, ratio of which is temperature dependent. Both isomers undergo in aqueous solution to spontaneous isomerization which does not occur when carbonato group is replaced by oxalato or malonato one. This suggests that the source of stereolability in these complexes is a strain imposed by the four-membered carbonato chelate ring. The equilibrium constants (K = fac/mer) increase in the order KGly < KVal < KAla. The total rate constants obtained at 45 °C and 55 °C increase in the order kAla < kVal < kGly. λ-mer-[CoCO3(en)(S)-Val)] isomer undergoes at 22 °C racemization at the octahedral centre yielding the mixture of 59% δ and 41% λ isomers.
Light Scattering of Particles Embedded in Soft Hydrogel
Authors: Tian Minmin , Tuzar Zdeněk , Munk Petr
Published: July 2002

This study is concerned with soft polyacrylamide gels in which are embedded either polystyrene latexes or block copolymer micelles with polystyrene cores and poly(methacrylic acid) shells. We have found that light scattering provides useful structure information about the systems. For polystyrene latexes, both molar mass and radius of gyration do not change during the embedding in the hydrogel; for block copolymer micelles, molar mass does not change, while the size decreases upon immobilization. In all cases, the second virial coefficient is very small and overshadowed by the turbidity of the sample. For large particles with narrow particle size distribution, a combination of Guinier and Zimm plots has been used for obtaining the particle size and molar mass.
Stress Whitening in Polypropylene. I. Light Scattering Theory and Model Experiments
Authors: Holoubek Jaroslav , Raab Miroslav
Published: July 2002

Theoretical background for an optical method is presented which makes it possible to distinguish unambiguously between voids and particles as light scattering sites in polymeric materials. Typical dependences of turbidity as a function of diameter of scattering elements, their volume fractions and also turbidity curves as a function of the wavelength of the incident light were calculated, based both on the Lorenz-Mie theory and the fluctuation theory. Such dependences calculated for polypropylene-containing voids on the one hand and particles, differing only slightly from the surrounding matrix in their refractive index, on the other hand, are markedly different. The most significant results are: (i) Turbidity is at least by two orders of magnitude larger for voids in comparison to embedded particles of ethylene-propylene (EPDM) rubber of the same size, concentration and at the same wavelength. (ii) The wavelength dependence of turbidity for EPDM particles and the inherent refractive index fluctuations in the polypropylene matrix is much steeper as compared to voids for all considered diameters (0.1-10 μm). Thus, the nature of stress whitening in complex polymeric materials can be determined from turbidity measurements.