bluuebunny/crossref_metadata_2025_split huggingface dataset:
Structural basis of α‐amylase activation by chloride
Authors: Aghajari Nushin , Feller Georges , Gerday Charles , Haser Richard 
Published: July 2002

AbstractTo further investigate the mechanism and function of allosteric activation by chloride in some α‐amylases, the structure of the bacterial α‐amylase from the psychrophilic micro‐organism Pseudoalteromonas haloplanktis in complex with nitrate has been solved at 2.1 A˚, as well as the structure of the mutants Lys300Gln (2.5 A˚) and Lys300Arg (2.25 A˚). Nitrate binds strongly to α‐amylase but is a weak activator. Mutation of the critical chloride ligand Lys300 into Gln results in a chloride‐independent enzyme, whereas the mutation into Arg mimics the binding site as is found in animal α‐amylases with, however, a lower affinity for chloride. These structures reveal that the triangular conformation of the chloride ligands and the nearly equatorial coordination allow the perfect accommodation of planar trigonal monovalent anions such as NO3−, explaining their unusual strong binding. It is also shown that a localized negative charge such as that of Cl−, rather than a delocalized charge as in the case of nitrate, is essential for maximal activation. The chloride‐free mutant Lys300Gln indicates that chloride is not mandatory for the catalytic mechanism but strongly increases the reactivity at the active site. Disappearance of the putative catalytic water molecule in this weakly active mutant supports the view that chloride helps to polarize the hydrolytic water molecule and enhances the rate of the second step in the catalytic reaction.
The Photometallic Process
Authors: Burgess J. F. , Schaefer D. L. 
Published: July 2002

The photometallic process is a new method of producing images in thin films of metals, semiconductors, and dielectrics. The process depends on the photogeneration of reactive material in situ which reacts with the thin film to produce a pattern.
A polymer layer containing a photodecomposable material is coated on a thin film. A pattern of light on the surface causes preferential photodecomposition and these products react with the thin film, removing it where exposed to light. The coating and reaction products are removed in a solvent rinse. A positive copy of the light image is thus obtained in the thin film. Since very close control of the photodecomposition is possible, images with continuous grey scale or various depths of reaction are possible. The process is exemplified by a description of the photoetching of gold, nichrome, and silicon dioxide. Other materials for which the process has been demonstrated are listed. A discussion of thin-film parameters, coating parameters, and proposed reaction mechanisms is presented. Among other applications, the process has potential use in microelectronic circuit fabrication.
Abstract: Pattern generation on wafers using the electron-beam exposure system (EBES)
Authors: Henderson R. C. , Voshchenkov A. M. , Mahoney G. E. 
Published: July 2002

Using the electron-beam exposure system (EBES) for direct pattern generation on wafers offers the means to make circuits with feature dimensions of only a few micrometers and alignment tolerances less than 1 μm. For such a program to be successful, techniques had to be developed to (a) rapidly register the pattern, (b) minimize the writing time, and (c) pattern the various thin films used in circuit fabrication.
For pattern registration, EBES need only determine the positions of three nonlinear fiducial marks which form a triangle spanning most of the wafer area. The position data are used to calculate the translation, rotation, skew, and magnification required to get the best fit of the old pattern to the new. The corrections are applied to the electron-beam writing operation. It takes less than two minutes to register an entire 2-in. (5.08-cm) wafer to within a ±1/2-μm accuracy.
The writing time is usually less than 20 min to expose all the chips fully covering a 2-in. (5.08-cm) Si wafer, even when there are three or four different types of circuit on the wafer. This can be achieved by laying out the chips to specifically match the EBES writing characteristics.
The resist used has been poly (glycidyl methacrylate-co-ethyl acrylate), a sensitive negative electron resist. Techniques were devised to etch patterns on many different substrates following resist development.
For pattern generation on Si wafers, the substrates included thin films of SiO2 (thermally grown as well as CVD), Si3N4, W, polysilicon, Al, and Ti–Pd–Au (see Table I). Subsequently, MOS circuits were made with combinations of the above techniques. These circuits2 include 1024-bit RAM’s, some with 5.5-μm gates, 3.0-μm contact holes, and 4.5-μm interconnecting metallization. Also, inverters, as well as discrete IGFET’s with gate lengths varying from 2 to 12 μm, were made.
EBES has also been used to delineate the thin film patterns required for GaAs MES FET fabrication (see Table II). Some devices have up to fourteen 2.5-μm gates with 2.5-μm gate-to-source spacings while others have single 2.0×500-μm gates with 2.0-μm spacings. These devices require alignment better than ±0.25 μm and metal conductors with thickness-to-width ratios as high as 1/2. Single gate MES FET’s made in this way have values of transconductance of up to 25 mmhos.
High-Level Expression of Chromosomally Encoded SHV-1 β-Lactamase and an Outer Membrane Protein Change Confer Resistance to Ceftazidime and Piperacillin- Tazobactam in a Clinical Isolate of
Klebsiella pneumoniae
Authors: Rice Louis B. , Carias Lenore L. , Hujer Andrea M. , Bonafede Mary , Hutton Rebecca , Hoyen Claudia , Bonomo Robert A. 
Published: July 2002

ABSTRACT

We describe
Klebsiella pneumoniae
15571, a clinical isolate resistant to ceftazidime MIC = 32 μg/ml) and piperacillin-tazobactam (MICs = 1,024 and 128 μg/ml).
K. pneumoniae
15571 expresses a single β-lactamase with a pI of 7.6. However, when cloned in a high-copy-number vector in
Escherichia coli
, this
bla
SHV-1
gene did not confer resistance to ceftazidime, a spectrum consistent with the nucleotide sequence, which was nearly identical to those of previously described
bla
SHV-1
genes. Outer membrane protein (OMP) analysis of
K. pneumoniae
15571 revealed a decrease in the quantity of a minor 45-kDa OMP in comparison to that in
K. pneumoniae
44NR, a low-level ampicillin-resistant strain that also expresses a chromosomally determined
bla
SHV-1
. Crude β-lactamase enzyme extracts from
K. pneumoniae
15571 produced roughly 200-fold more β-lactamase activity than
K. pneumoniae
44NR. Northern hybridization analysis revealed that this difference was explainable by quantifiable differences in transcription of the
bla
SHV-1
gene in the two strains. Primer extension analysis of
bla
SHV-1
mRNA from
K. pneumoniae
15571 and 44NR indicated that the transcriptional start sites were identical in the two strains. DNA sequencing of the promoter regions upstream of the of
bla
SHV-1
open reading frames in the two
K. pneumoniae
strains revealed an A→C change in the second position of the −10 region in
K. pneumoniae
44NR compared to that in 15571. Site-directed mutagenesis of the cloned
K. pneumoniae
15571
bla
SHV-1
, in which the A in the second position of the 15571 −10 region was changed to a C, resulted in a substantial lowering of the MIC of ampicillin. When the levels of β-lactamase enzyme expression in
E. coli
were compared, the
bla
SHV-1
downstream of the altered −10 region produced 17-fold less β-lactamase enzyme. These results indicate that elevated levels of ceftazidime resistance can result from a combination of increased enzyme production and minor OMP changes and that levels of chromosomally encoded SHV-1 β-lactamase production can vary substantially with a single-base-pair change in promoter sequence.
Pharmacokinetics and Pharmacodynamics of Intravenous Artesunate in Severe Falciparum Malaria
Authors: Davis Timothy M. E. , Phuong Hoang Lan , Ilett Kenneth F. , Hung Nguyen Canh , Batty Kevin T. , Phuong Vu Duong Bich , Powell Shane M. , Thien Huynh Van , Binh Tran Quang 
Published: July 2002

ABSTRACT

To provide novel data relating to the dispositions, effects, and toxicities of the artemisinin derivatives in severe malaria, we studied 30 Vietnamese adults with slide-positive falciparum malaria treated with intravenous artesunate. Twelve patients with complications (severe; group 1) and 8 patients without complications but requiring parenteral therapy (moderately severe; group 2) received 120 mg of artesunate by injection, and 10 patients with moderately severe complications (group 3) were given 240 mg by infusion. Serial concentrations of artesunate and its active metabolite dihydroartemisinin in plasma were measured by high-performance liquid chromatography. The time to 50% parasite clearance (PCT
50
) was determined from serial parasite densities. Full clinical (including neurological) assessments were performed at least daily. In noncompartmental pharmacokinetic analyses, group mean artesunate half-lives (
t
1/2
) were short (range, 2.3 to 4.3 min). The dihydroartemisinin
t
1/2
(range, 40 to 64 min), clearance (range, 0.73 to 1.01 liters/h/kg), and volume of distribution (range, 0.77 to 1.01 liters/kg) were also similar both across the three patient groups (
P
> 0.1) and to previously reported values for patients with uncomplicated malaria. Parasite clearance was prompt (group median PCT
50
range 6 to 9 h) and clinical recovery was complete under all three regimens. These data indicate that the pharmacokinetics of artesunate and dihydroartemisinin are not influenced by the severity of malaria. Since the pharmacokinetic parameters for both artesunate and dihydroartemisinin were similar regardless of whether injection or infusion was used, artesunate can be considered a prodrug that is converted stoichiometrically to dhydroartemisinin. Conventional doses of artesunate are safe and effective when given to patients with complications of falciparum malaria.
In Vitro Activities of Two Antimitotic Compounds, Pancratistatin and 7-Deoxynarciclasine, against
Encephalitozoon intestinalis
, a Microsporidium Causing Infections in Humans
Authors: Ouarzane-Amara Meryem , Franetich Jean-François , Mazier Dominique , Pettit George R. , Meijer Laurent , Doerig Christian , Desportes-Livage Isabelle 
Published: July 2002

ABSTRACT

The antiparasitic effect of a collection of compounds with antimitotic activity has been tested on a mammalian cell line infected with
Encephalitozoon intestinalis
, a microsporidian causing intestinal and systemic infection in immunocompromised patients. The antiparasitic effect was evaluated by counting the number of parasitophorous vacuoles detected by immunofluorescence. Out of 526 compounds tested, 2 (pancratistatin and 7-deoxynarciclasine) inhibited the infection without affecting the host cell. The 50% inhibitory concentrations (IC
50
s) of pancratistatin and 7-deoxynarciclasine for
E. intestinalis
were 0.18 μM and 0.2 μM, respectively, approximately eightfold lower than the IC
50
s of these same compounds against the host cells. Electron microscopy confirmed the gradual decrease in the number of parasitophorous vacuoles and showed that of the two life cycle phases, sporogony was more sensitive to the inhibitors than merogony. Furthermore, the persistence of meronts in some cells apparently devoid of sporonts and spores indicated that the inhibitors block development rather than entry of the parasite into the host cell. The occurrence of binucleate sporoblasts and spores suggests that these inhibitors blocked a specific phase of cell division.
Characterization of Mutations in DNA Gyrase and Topoisomerase IV Involved in Quinolone Resistance of
Mycoplasma gallisepticum
Mutants Obtained In Vitro
Authors: Reinhardt A. K. , Bébéar C. M. , Kobisch M. , Kempf I. , Gautier-Bouchardon A. V. 
Published: July 2002

ABSTRACT

Mycoplasma gallisepticum
enrofloxacin-resistant mutants were generated by stepwise selection in increasing concentrations of enrofloxacin. Alterations were found in the quinolone resistance-determining regions of the four target genes encoding DNA gyrase and topoisomerase IV from these mutants. This is the first description of such mutations in an animal mycoplasma species.
Molecular Characterization of a Novel Class 1 Integron Containing
bla
GES-1
and a Fused Product of
aac(3)-Ib/aac(6
"
)-Ib
" Gene Cassettes in
Pseudomonas aeruginosa
Authors: Dubois Véronique , Poirel Laurent , Marie Caroline , Arpin Corinne , Nordmann Patrice , Quentin Claudine 
Published: July 2002

ABSTRACT

As seen by the disk diffusion method, the clinical strain of
Pseudomonas aeruginosa
Pa695, resistant to all extended-spectrum cephalosporins and aminoglycosides, exhibited an unusual synergistic effect between ceftazidime and imipenem. This isolate produced an extended-spectrum β-lactamase (ESBL) with a pI of 5.8 that appeared to be chromosomally encoded. Cloning experiments revealed that this ESBL was encoded by
bla
GES-1
, previously described in an integron from
Klebsiella pneumoniae
. In
P. aeruginosa
Pa695, a higher level of resistance to ceftazidime than to ticarcillin was observed, and no synergy between the β-lactamase inhibitors and extended-spectrum cephalosporins was detected, in contrast to the resistance pattern observed in
K. pneumoniae
. Further sequence analysis demonstrated that the
bla
GES-1
gene cassette was located in a class 1 integron, which contained another sequence corresponding to the fused
aac(3)-Ib
and
aac(6
"
)-Ib
" gene cassettes. The fusion product was functional, as was the product of each gene cloned separately: AAC(3)-I, despite the deletion of the four last amino acids, and AAC(6"), which carried three amino acid changes compared with the most homologous sequence. The AAC(3)-I protein conferred an expected gentamicin and fortimicin resistance, and the AAC(6"), despite the Leu-119→Ser substitution, yielded resistance to kanamycin, tobramycin, and dibekacin, but slightly affected netilmicin and amikacin, and had no apparent effect on gentamicin. The fusion product conveyed a large profile of resistance, combining the AAC(6") activity with a higher level of gentamicin resistance without accompanying fortimicin resistance.
Impact of
N
-Chlorotaurine on Viability and Production of Secreted Aspartyl Proteinases of
Candida
spp
Authors: Nagl Markus , Gruber Andreas , Fuchs Anita , Lell Claudia P. , Lemberger Eva-Maria , Borg-von Zepelin Margarete , Würzner Reinhard 
Published: July 2002

ABSTRACT

N
-Chlorotaurine, an endogenous long-lived oxidant, demonstrated fungicidal activity against
Candida
spp. and a postantifungal effect. Secreted aspartyl proteinases, important fungal virulence factors, proved to be a first target of impact. These results provide support for the topical application of
N
-chlorotaurine as an antimicrobial agent in yeast infections.
A Strain of
Enterococcus faecium
(18C23) Inhibits Adhesion of Enterotoxigenic
Escherichia coli
K88 to Porcine Small Intestine Mucus
Authors: Jin L. Z. , Marquardt R. R. , Zhao X. 
Published: July 2002

ABSTRACT

Few studies, if any, have addressed the adhesion of enterococci to the intestinal mucosa and their interference with the adhesion of pathogens, although more than 60% of probiotic preparations in the market contain strains of enterococci. The objective of this study was to investigate if
Enterococcus faecium
18C23 has the ability to inhibit the adhesion of
Escherichia coli
K88ac and K88MB to the small intestine mucus of piglets. Approximately 9% of
E. faecium
18C23 organisms adhered to the small intestine mucus, and the adhesion was found to be specific. Living
E. faecium
18C23 culture efficiently inhibited the adhesion of
E. coli
K88ac and K88MB to the piglet intestine mucus. Inhibition of the adhesion of
E. coli
K88ac to the small intestine mucus was found to be dose dependent. Inhibition of >90% was observed when 10
9
CFU or more of living
E. faecium
18C23 culture per ml was added simultaneously with
E. coli
to immobilized mucus. The substances from both the 18C23 cells and the spent culture supernatant contributed to the inhibition of adhesion of
E. coli
K88 to the small intestine mucus receptors. The inhibiting effect was not solely a pH effect since considerable inhibitory action was demonstrated after neutralizing the mixture or spent culture supernatant to pH 7.0. Part of the inhibition of adhesion of
E. coli
K88ac by
E. faecium
18C23 or its supernatant might occur through steric hindrance.
Influence of Catalase and Superoxide Dismutase on Ozone Inactivation of
Listeria monocytogenes
Authors: Fisher Christopher W. , Lee Dongha , Dodge Beth-Anne , Hamman Kristen M. , Robbins Justin B. , Martin Scott E. 
Published: July 2002

ABSTRACT

The effects of ozone at 0.25, 0.40, and 1.00 ppm on
Listeria monocytogenes
were evaluated in distilled water and phosphate-buffered saline. Differences in sensitivity to ozone were found to exist among the six strains examined. Greater cell death was found following exposure at lower temperatures. Early stationary-phase cells were less sensitive to ozone than mid-exponential- and late stationary-phase cells. Ozonation at 1.00 ppm of cabbage inoculated with
L. monocytogenes
effectively inactivated all cells after 5 min. The abilities of in vivo catalase and superoxide dismutase to protect the cells from ozone were also examined. Three listerial test strains were inactivated rapidly upon exposure to ozone. Both catalase and superoxide dismutase were found to protect listerial cells from ozone attack, with superoxide dismutase being more important than catalase in this protection.
Chlorine-Susceptible and Chlorine-Resistant Type 021N Bacteria Occurring in Bulking Activated Sludges
Authors: Séka M. A. , Kalogo Y. , Hammes F. , Kielemoes J. , Verstraete W. 
Published: July 2002

ABSTRACT
Two filamentous bacteria causing bulking in two activated sludges were examined. Investigations using morphological features, staining techniques, and fluorescent in situ hybridization identified both filaments as type 021N. However, an examination of the effect of chlorine on the sludges revealed a chlorine-susceptible type 021N in one sludge and a chlorine-resistant type 021N in the other.
Chromosomal Gene Inactivation in the Green Sulfur Bacterium
Chlorobium tepidum
by Natural Transformation
Authors: Frigaard Niels-Ulrik , Bryant Donald A. 
Published: July 2002

ABSTRACT

Conditions for inactivating chromosomal genes of
Chlorobium tepidum
by natural transformation and homologous recombination were established. As a model, mutants unable to perform nitrogen fixation were constructed by interrupting
nifD
with various antibiotic resistance markers. Growth of wild-type
C. tepidum
at 40°C on agar plates could be completely inhibited by 100 μg of gentamicin ml
−1
, 2 μg of erythromycin ml
−1
, 30 μg of chloramphenicol ml
−1
, or 1 μg of tetracycline ml
−1
or a combination of 300 μg of streptomycin ml
−1
and 150 μg of spectinomycin ml
−1
. Transformation was performed by spotting cells and DNA on an agar plate for 10 to 20 h. Transformation frequencies on the order of 10
−7
were observed with gentamicin and erythromycin markers, and transformation frequencies on the order of 10
−3
were observed with a streptomycin-spectinomycin marker. The frequency of spontaneous mutants resistant to gentamicin, erythromycin, or spectinomycin-streptomycin was undetectable or significantly lower than the transformation frequency. Transformation with the gentamicin marker was observed when the transforming DNA contained 1 or 3 kb of total homologous flanking sequence but not when the transforming DNA contained only 0.3 kb of homologous sequence. Linearized plasmids transformed at least an order of magnitude better than circular plasmids. This work forms a foundation for the systematic targeted inactivation of genes in
C. tepidum
, whose 2.15-Mb genome has recently been completely sequenced.
Antacid Increases Survival of
Vibrio vulnificus
and
Vibrio vulnificus
Phage in a Gastrointestinal Model
Authors: Koo Jaheon , Marshall Douglas L. , DePaola Angelo 
Published: July 2002

ABSTRACT

Viable counts of three strains of
Vibrio vulnificus
and its phage were determined during exposure to a mechanical gastrointestinal model with or without antacid for 9 h at 37°C.
V. vulnificus
was eliminated (>4-log reduction) within 30 min in the gastric compartment (pH decline from 5.0 to 3.5). Viable
V. vulnificus
cells delivered from the gastric compartment during the first 30 min of exposure reached 10
6
to 10
8
CFU/ml in the intestinal compartment after 9 h (pH 7.0). Phages were eliminated within 45 min in the gastric compartment (pH decline from 5.1 to 2.5). Less than a 2-log reduction of phage was observed in the intestinal compartment after 9 h (pH 7.0). When the gastric compartment contained antacid
V. vulnificus
counts decreased slightly (<2 log) during 2 h of exposure (pH decline from 7.7 to 6.0), while counts in the intestinal compartment (pH 7.5) reached 10
7
to 10
9
CFU/ml. Phage numbers decreased 1 log after 2 h in the gastric compartment (pH decline from 7.7 to 5.7) containing antacid and decreased 1 log in the intestinal compartment (pH 7.6) after 9 h. Presence of antacid in the gastric compartment of the model greatly increased the ability of both
V. vulnificus
and its phage to survive simulated gastrointestinal transit and may be a factor involved with oyster-associated illness.
Sinorhizobium meliloti
Cells Require Biotin and either Cobalt or Methionine for Growth
Authors: Watson Robert J. , Heys Roselyn , Martin Teresa , Savard Marc 
Published: July 2002

ABSTRACT

Sinorhizobium meliloti
is usually cultured in rich media containing yeast extract. It has been suggested that some components of yeast extract are also required for growth in minimal medium. We tested 27 strains of this bacterium and found that none were able to grow in minimal medium when methods to limit carryover of yeast extract were used during inoculation. By fractionation of yeast extract, two required growth factors were identified. Biotin was found to be absolutely required for growth, whereas previously the need for this vitamin was considered to be strain specific. All strains also required supplementation with cobalt or methionine, consistent with the requirement for a vitamin B
12
-dependent homocysteine methyltransferase for methionine biosynthesis.
Isomaltulose Synthase from
Klebsiella
sp. Strain LX3: Gene Cloning and Characterization and Engineering of Thermostability
Authors: Zhang Daohai , Li Xianzhen , Zhang Lian-Hui 
Published: July 2002

ABSTRACT

The gene (
palI
) encoding isomaltulose synthase (PalI) from a soil bacterial isolate,
Klebsiella
sp. strain LX3, was cloned and characterized. PalI converts sucrose into isomaltulose, trehalulose, and trace amounts of glucose and fructose. Sequence domain analysis showed that PalI contains an α-amylase domain and (β/α)
8
-barrel structures, suggesting that it belongs to the α-amylase family. Sequence alignment indicated that the five amino acid residues of catalytic importance in α-amylases and glucosyltransferases (Asp
241
, Glu
295
, Asp
369
, His
145
, and His
368
) are conserved in PalI. Purified recombinant PalI displayed high catalytic efficiency, with a

K
m

of 54.6 ± 1.7 mM for sucrose, and maximum activity (approximately 328.0 ± 2.5 U/mg) at pH 6.0 and 35°C. PalI activity was strongly inhibited by Fe
3+
and Hg
2+
and was enhanced by Mn
2+
and Mg
2+
. The half-life of PalI was 1.8 min at 50°C. Replacement of selected amino acid residues by proline significantly increased the thermostability of PalI. Simultaneous replacement of Glu
498
and Arg
310
with proline resulted in an 11-fold increase in the half-life of PalI at 50°C.