bluuebunny/crossref_metadata_2025_split huggingface dataset:
Brassinosteroid-6-Oxidases from Arabidopsis and Tomato Catalyze Multiple C-6 Oxidations in Brassinosteroid Biosynthesis
Authors: Shimada Yukihisa , Fujioka Shozo , Miyauchi Narumasa , Kushiro Masayo , Takatsuto Suguru , Nomura Takahito , Yokota Takao , Kamiya Yuji , Bishop Gerard J. , Yoshida Shigeo 
Published: July 2002

AbstractBrassinosteroids (BRs) are steroidal plant hormones that are essential for growth and development. It has been proposed that BRs are synthesized via two parallel pathways, the early and late C-6 oxidation pathways according to the C-6 oxidation status. The tomato (Lycopersicon esculentum) Dwarf gene encodes a cytochrome P450 that has been shown to catalyze the C-6 oxidation of 6-deoxocastasterone to castasterone. We isolated an Arabidopsis ortholog (AtBR6ox gene) of the tomatoDwarf gene. The encoded polypeptide has characteristics of P450s and is classified into the CYP85 family. TheAtBR6ox and tomato Dwarf gene were expressed in yeast and the ability of the transformed yeast cells to metabolize 6-deoxo-BRs was tested. Metabolites were analyzed by gas chromatography-mass spectrometry. Both enzymes catalyze multiple steps in BR biosynthesis: 6-deoxoteasterone to teasterone, 3-dehydro-6-deoxoteasterone to 3-dehydroteasterone, 6-deoxotyphasterol to typhasterol, and 6-deoxocastasterone to castasterone. Our results indicate that the AtBR6ox gene and the tomatoDwarf gene encode steroid-6-oxidases and that these enzymes have a broad substrate specificity. This suggests that the BR biosynthetic pathway consists of a metabolic grid rather than two separate parallel pathways.
The Wheat cDNA LCT1 Generates Hypersensitivity to Sodium in a Salt-Sensitive Yeast Strain
Authors: Amtmann Anna , Fischer Marc , Marsh Ellen L. , Stefanovic Aleksandra , Sanders Dale , Schachtman Daniel P. 
Published: July 2002

Abstract
Salinity affects large areas of agricultural land, and all major crop species are intolerant to high levels of sodium ions. The principal route for Na+ uptake into plant cells remains to be identified. Non-selective ion channels and high-affinity potassium transporters have emerged as potential pathways for Na+entry. A third candidate for Na+ transport into plant cells is a low-affinity cation transporter represented by the wheat protein LCT1, which is known to be permeable for a wide range of cations when expressed in yeast (Saccharomyces cerevisiae). To investigate the role of LCT1 in salt tolerance we have used the yeast strain G19, which is disrupted in the genes encoding Na+export pumps and as a result displays salt sensitivity comparable with wheat. After transformation with LCT1, G19 cells became hypersensitive to NaCl. We show that LCT1 expression results in a strong decrease of intracellular K+/Na+ ratio in G19 cells due to the combined effect of enhanced Na+ accumulation and loss of intracellular K+. Na+ uptake through LCT1 was inhibited by K+ and Ca2+ at high concentrations and the addition of these ions rescued growth ofLCT1-transformed G19 on saline medium. LCT1 was also shown to mediate the uptake of Li+ and Cs+. Expression of two mutant LCT1 cDNAs with N-terminal truncations resulted in decreased Ca2+ uptake and increased Na+ tolerance compared with expression of the full-lengthLCT1. Our findings strongly suggest thatLCT1 represents a molecular link between Ca2+ and Na+ uptake into plant cells.
A model of the replication fork blocking protein Fob1p based on the catalytic core domain of retroviral integrases
Authors: Dlakić Mensur 
Published: July 2002

AbstractThe replication fork blocks are common in both prokaryotes and eukaryotes. In most cases, these blocks are associated with increased levels of mitotic recombination. One of the best‐characterized replication fork blocks in eukaryotes is found in ribosomal DNA (rDNA) repeats of Saccharomyces cerevisiae. It has been shown that the replication fork blocking protein Fob1p regulates the recombination rate and the number of rDNA copies in S. cerevisiae, but the mechanistic aspects of these events are still poorly understood. Sequence profile searches revealed that Fob1p is related to retroviral integrases. Subsequently, the catalytic domain of HIV‐1 integrase was used as a template to build a reliable three‐dimensional model of Fob1p. Structural insights from this study may be useful in explaining Fob1p‐mediated formation of extrachromosomal rDNA circles that accelerate aging in yeast and recombination events that lead to expansion or contraction of rDNA.
Distance dependence and salt sensitivity of pairwise, coulombic interactions in a protein
Authors: Lee Kelly K. , Fitch Carolyn A. , García‐Moreno E. Bertrand 
Published: July 2002

AbstractHistidine pKa values were measured in charge‐reversal (K78E, K97E, K127E, and K97E/K127E) and charge‐neutralization (E10A, E101A, and R35A) mutants of staphylococcal nuclease (SNase) by 1H‐NMR spectroscopy. Energies of interaction between pairs of charges (ΔGij) were obtained from the shifts in pKa values relative to wild‐type values. The data describe the distance dependence and salt sensitivity of pairwise coulombic interactions. Calculations with a continuum electrostatics method captured the experimental ΔGij when static structures were used and when the protein interior was treated empirically with a dielectric constant of 20. The ΔGij when rij ≤ 10 Å were exaggerated slightly in the calculations. Coulomb's law with a dielectric constant near 80 and a Debye‐Hückel term to account for screening by the ionic strength reproduced the salt sensitivity and distance dependence of ΔGij as well as the structure‐based method. In their interactions with each other, surface charges behave as if immersed in water; the Debye length describes realistically the distance where interactions become negligible at a given ionic strength. On average, charges separated by distances (rij) ≈5 Å interacted with ΔGij ≈ 0.6 kcal/mole in 0.01 M KCl, but ΔGij decayed to ≤0.10 kcal/mole when rij = 20 Å. In 0.10 M KCl, ΔGij ≈ 0.10 kcal/mole when rij = 10 Å. In 1.5 M KCl, only short‐range interactions with rij ≤ 5 Å persisted. Although at physiological ionic strengths the interactions between charges separated by more than 10 Å are extremely weak, in situations where charge imbalance exists many weak interactions can cumulatively produce substantial effects.
Vacuum Problems of Electron and Positron Storage Rings
Authors: Bernardini M. , Malter L. 
Published: July 2002

The problem of gas evolution in electron-positron storage rings due to synchrotron radiation was explored. The spectral distribution of the radiation was first computed. From this and from data on quantum efficiency it was possible to determine an upper limit to the resultant photoemission from the ring walls. Measurements of the desorption efficiency from stainless steel 304 then made possible a determination of the maximum gas load which could be expected as a result of the synchrotron radiation.
This treatment was applied to a planned storage ring to be built at Frascati. From this it appeared that with modest processing the ring walls could be put into a condition such that the gas evolution would have completely acceptable values. The effects of wall temperature and of processing in hydrogen and oxygen upon the desorption efficiency were determined. On the basis of the results obtained it is concluded that optimum results would be obtained by the use of materials which are gas-free and on which tightly bound layers are formed from the ambient atmosphere.
Bactericidal Activity of Quinupristin-Dalfopristin against
Staphylococcus aureus
: Clindamycin Susceptibility as a Surrogate Indicator
Authors: Fuchs Peter C. , Barry Arthur L. , Brown Steven D. 
Published: July 2002

ABSTRACT

Of 516
Staphylococcus aureus
strains tested, 97.1% were susceptible to quinupristin-dalfopristin, which was bactericidal for 22 (56%) of the 39 strains tested, comparable to vancomycin. All 17 clindamycin and macrolide-resistant strains were inhibited but not killed by quinupristin-dalfopristin, whereas all 22 clindamycin-susceptible strains (5 were macrolide resistant) were killed.
Intravitreal, Retinal, and Central Nervous System Foscarnet Concentrations after Rapid Intravenous Administration to Rabbits
Authors: López-Cortés Luis F. , Ruiz-Valderas R. , Lucero-Muñoz M. J. , Cordero E. , Pastor-Ramos M. T. , Marquez J. 
Published: July 2002

ABSTRACT
Retinal, vitreous humor, brain, and cerebrospinal fluid (CSF) foscarnet levels were measured by high-performance liquid chromatography after administration of an intravenous dose of 120 mg/kg of body weight to 32 pigmented rabbits. A pharmacokinetic analysis was done using a two-compartment model. The penetration ratios, defined as ratios of retinal, vitreous humor, brain, and CSF areas under the concentration-time curve from 0 to 2 h were 110% ± 1%, 12.3% ± 0.7%, 118% ± 1%, and 20.2% ± 2.2%, respectively. These results suggest a good penetration of foscarnet into the retinal and brain tissues, reaching higher concentrations than those estimated from vitreous humor and CSF levels.
Selective Interaction of the Human Immunodeficiency Virus Type 1 Reverse Transcriptase Nonnucleoside Inhibitor Efavirenz and Its Thio-Substituted Analog with Different Enzyme-Substrate Complexes
Authors: Maga Giovanni , Ubiali Daniela , Salvetti Raul , Pregnolato Massimo , Spadari Silvio 
Published: July 2002

ABSTRACT

Accumulating data have brought the nonnucleoside reverse transcriptase (RT) inhibitors (NNRTIs) into the forefront of antiretroviral therapy. Among the emerging compounds in this class, a particularly attractive one is efavirenz (Sustiva), recently approved for clinical use by the U.S. Food and Drug Administration. In the present study, the equilibrium dissociation constants for efavirenz binding to the different catalytic forms of human immunodeficiency virus type 1 RT as well as the association and dissociation rates have been determined using a steady-state kinetic approach. In addition, the same enzymological analysis has been extended to the thio-substituted analog, sefavirenz, which showed comparable activity in vitro against RT. Both compounds have been found to act as purely uncompetitive inhibitors at low drug concentrations (5 to 50 nM) and as mixed noncompetitive inhibitors at higher doses (50 to 500 nM). This behavior can be interpreted in terms of the relative affinities for the different catalytic forms of the enzyme. Both efavirenz and sefavirenz showed increasing affinities for the different forms of RT in the following order: free enzyme < (i.e., bound with lower affinity) binary RT–template-primer (TP) complex < ternary RT-TP-deoxynucleoside triphosphate (dNTP) complex. The rate of binding of the two inhibitors to the different enzyme-substrate complexes was well below the diffusion limit (on the order of 10
4
M
−1
s
−1
); however, both inhibitors, when bound to the ternary RT-TP-dNTP complex, showed very low dissociation rates, on the order of 10
−4
s
−1
for both compounds, typical of tightly binding inhibitors. Thus, efavirenz and its thio-substituted derivative sefavirenz appear to be peculiar in their mechanism of action, being selective tightly binding inhibitors of the ternary RT-TP-dNTP complex. Efavirenz is the first clinically approved NNRTI to show this property.
Antimicrobial Susceptibility and Clinical Sources of
Dolosigranulum pigrum
Cultures
Authors: Laclaire L. , Facklam R. 
Published: July 2002

ABSTRACT

Antimicrobial susceptibilities of 27 clinical isolates of
Dolosigranulum pigrum
were determined. All were susceptible to amoxicillin, cefotaxime, cefuroxime, clindamycin, levofloxacin, meropenem, penicillin, quinupristin-dalfopristin, rifampin, tetracycline, and vancomycin. Fifteen of the isolates were intermediate to chloramphenicol. One isolate was resistant to trimethoprim-sulfamethoxazole. Two isolates were susceptible, 10 were intermediate, and 15 were resistant to erythromycin.
Identification and Characterization of
vanB2
Glycopeptide Resistance Elements in Enterococci Isolated in Scotland
Authors: McGregor K. F. , Young H.-K. 
Published: July 2002

ABSTRACT

Thirty-two
vanB
glycopeptide-resistant enterococci (28
Enterococcus faecium
and 4
Enterococcus faecalis
) were collected from hospitalized patients in Glasgow, Edinburgh, Dundee, and Aberdeen, Scotland, and the
vanB
element in each was compared to
vanB1
of
E. faecalis
strain ATCC 51299.
Hha
I digestion of PCR fragments of the
vanB
ligase gene was used to identify
vanB
subtypes. All
E. faecium
isolates were
vanB2
, and all
E. faecalis
isolates were
vanB1
. Restriction fragment length polymorphism analysis of a 5,180-bp

vanS
B
-vanX
B

long-PCR fragment of the
vanB
cluster showed the loss of
Hae
II restriction sites in

vanS
B

,
vanW
, and

vanX
B

in strains containing a
vanB2
ligase gene. Partial sequences of genes in the
vanB2
cluster for two genomically distinct Scottish isolates were >99.8% identical to each other.

vanS
B2

,

vanX
B2

, and
vanB2
sequences differed at the nucleotide level from those of

vanS
B

,

vanX
B

, and
vanB
by 4.2, 4.6, and 4.8%, respectively. The
vanB2
resistance element appears to be widespread among VanB glycopeptide-resistant
E. faecium
strains isolated in Scottish hospitals.
Comparative In Vitro Activities of Ertapenem (MK-0826) against 1,001 Anaerobes Isolated from Human Intra-Abdominal Infections
Authors: Goldstein Ellie J. C. , Citron Diane M. , Vreni Merriam C. , Warren Yumi , Tyrrell Kerin L. 
Published: July 2002

ABSTRACT

By using an agar dilution method, the comparative in vitro activities of ertapenem (MK-0826) were studied against 1,001 anaerobes isolated from human intra-abdominal infections in 17 countries worldwide. MK-0826 was uniformly active against all isolates, including all
Bacteroides fragilis
group species isolates, with the exception of 12 of 61 (20%) strains of
Bilophila wadsworthia
, 3 strains of lactobacilli, and 1 isolate of
Acidaminococcus fermentans
. Geographical variation in activity was not observed.
Antimalarial Activities of Aminooxy Compounds
Authors: Berger Bradley J. 
Published: July 2002

ABSTRACT

Twenty-three aminooxy compounds have been examined for their ability to inhibit the growth of the malaria parasite
Plasmodium falciparum
in vitro. Eight of these compounds were found to have 50% inhibitory concentrations less than 10 μM, with the best drugs being canaline (the aminooxy analogue of ornithine) and CGP51905A at 297 ± 23.6 nM and 242 ± 18.8 nM, respectively. Canaline was also assayed in combination with the ornithine decarboxylase inhibitor difluoromethylornithine, and the two drugs were found to be synergistic in antimalarial activity.
Evaluation of Current Activities of Fluoroquinolones against Gram-Negative Bacilli Using Centralized In Vitro Testing and Electronic Surveillance
Authors: Sahm Daniel F. , Critchley Ian A. , Kelly Laurie J. , Karlowsky James A. , Mayfield David C. , Thornsberry Clyde , Mauriz Yolanda R. , Kahn James 
Published: July 2002

ABSTRACT

Given the propensity for
Enterobacteriaceae
and clinically significant nonfermentative gram-negative bacilli to acquire antimicrobial resistance, consistent surveillance of the activities of agents commonly prescribed to treat infections arising from these organisms is imperative. This study determined the activities of two fluoroquinolones, levofloxacin and ciprofloxacin, and seven comparative agents against recent clinical isolates of
Enterobacteriaceae
,
Pseudomonas aeruginosa
,
Acinetobacter baumannii
, and
Stenotrophomonas maltophilia
using two surveillance strategies: 1) centralized in vitro susceptibility testing of isolates collected from 27 hospital laboratories across the United States and 2) analysis of data from The Surveillance Network Database-USA, an electronic surveillance network comprising more than 200 laboratories nationwide. Regardless of the surveillance method,
Enterobacteriaceae
,
P. aeruginosa
, and
A. baumannii
demonstrated similar rates of susceptibility to levofloxacin and ciprofloxacin. Susceptibilities to the fluoroquinolones approached or exceeded 90% for all
Enterobacteriaceae
except
Providencia
spp. (≤65%). Approximately 70% of
P. aeruginosa
and 50% of
A. baumanii
isolates were susceptible to both fluoroquinolones. Among
S. maltophilia
isolates, 50% more isolates were susceptible to levofloxacin than to ciprofloxacin. Overall, the rate of ceftazidime nonsusceptibility among
Enterobacteriaceae
was 8.7%, with fluoroquinolone resistance rates notably higher among ceftazidime-nonsusceptible isolates than ceftazidime-susceptible ones. Multidrug-resistant isolates were present among all species tested but were most prevalent for
Klebsiella pneumoniae
and
Enterobacter cloacae
. No gram-negative isolates resistant only to a fluoroquinolone were encountered, regardless of species. Thus, while levofloxacin and ciprofloxacin have maintained potent activity against
Enterobacteriaceae
, the potential for fluoroquinolone resistance, the apparent association between fluoroquinolone and cephalosporin resistance, and the presence of multidrug resistance in every species examined emphasize the need to maintain active surveillance of resistance patterns among gram-negative bacilli.
Sequencing of Cytomegalovirus UL97 Gene for Genotypic Antiviral Resistance Testing
Authors: Lurain Nell S. , Weinberg Adriana , Crumpacker Clyde S. , Chou Sunwen 
Published: July 2002

ABSTRACT
The widespread use of ganciclovir (GCV) to treat cytomegalovirus (CMV) infections in immunosuppressed patients has led to the development of drug resistance. Phenotypic assays for CMV drug resistance are presently too time-consuming to be therapeutically useful. To support the development of genotypic assays for GCV resistance, the complete sequences of the UL97 phosphotransferase genes in 28 phenotypically GCV-sensitive CMV clinical isolates were determined. The gene was found to be highly conserved, with nucleotide sequence identity among strains ranging from 98.6 to 100% and amino acid sequence identity of >99%. Primers for a genotypic assay were designed to amplify codons 400 to 707, because all known UL97 mutations conferring drug resistance occur at three sites within this region. This part of the UL97 gene was amplified from over 50 clinical isolates, and two sequencing reactions for the coding strand were successfully used to identify GCV resistance mutations. This genotypic assay can be performed in 48 h using genomic DNA extracted from cell monolayers at very low levels of virus infectivity, thus rapidly providing therapeutically useful results.
Measurement of Effects of Antibiotics in Bioluminescent
Staphylococcus aureus
RN4220
Authors: Tenhami Mervi , Hakkila Kaisa , Karp Matti 
Published: July 2002

ABSTRACT

The spread of antibiotic resistance among pathogenic bacteria is a serious threat to humans and animals. Therefore, unnecessary use should be minimized, and new antimicrobial agents with novel mechanisms of action are needed. We have developed an efficient method for measuring the action of antibiotics which is applied to a gram-positive strain,
Staphylococcus aureus
RN4220. The method utilizes the firefly luciferase reporter gene coupled to the metal-inducible
cadA
promoter in a plasmid, pTOO24. Correctly timed induction by micromolar concentrations of antimonite rapidly triggers the luciferase gene transcription and translation. This sensitizes the detection system to the action of antibiotics, and especially for transcriptional and translational inhibitors. We show the results for 11 model antibiotics with the present approach and compare them to an analytical setup with a strain where luciferase expression is under the regulation of a constitutive promoter giving only a report of metabolic inhibition. The measurement of light emission from intact living cells is shown to correlate extremely well (
r
= 0.99) with the conventional overnight growth inhibition measurement. Four of the antibiotics were within a 20% concentration range and four were within a 60% concentration range of the drugs tested. This approach shortens the assay time needed, and it can be performed in 1 to 4 h, depending on the sensitivity needed. Furthermore, the assay can be automatized for high-throughput screening by the pharmaceutical industry.
Carbapenem Resistance Mechanisms in
Pseudomonas aeruginosa
Clinical Isolates
Authors: Pai Hyunjoo , Kim Jong-Won , Kim Jungmin , Lee Ji Hyang , Choe Kang Won , Gotoh Naomasa 
Published: July 2002

ABSTRACT

In order to define the contributions of the mechanisms for carbapenem resistance in clinical strains of
Pseudomonas aeruginosa
, we investigated the presence of OprD, the expressions of the MexAB-OprM and MexEF-OprN systems, and the production of the β-lactamases for 44 clinical strains. All of the carbapenem-resistant isolates showed the loss of or decreased levels of OprD. Three strains overexpressed the MexAB-OprM efflux system by carrying mutations in
mexR
. These three strains had the amino acid substitution in MexR protein, Arg (CGG) → Gln (CAG), at the position of amino acid 70. None of the isolates, however, expressed the MexEF-OprN efflux system. For the characterization of β-lactamases, at least 13 isolates were the depressed mutants, and 12 strains produced secondary β-lactamases. Based on the above resistance mechanisms, the MICs of carbapenem for the isolates were analyzed. The MICs of carbapenem were mostly determined by the expression of OprD. The MICs of meropenem were two- to four-fold increased for the isolates which overexpressed MexAB-OprM in the background of OprD loss. However, the elevated MICs of meropenem for some individual isolates could not be explained. These findings suggested that other resistance mechanisms would play a role in meropenem resistance in clinical isolates of
P. aeruginosa
.
Characterization of Class 1 Integrons from
Pseudomonas aeruginosa
That Contain the
bla
VIM-2
Carbapenem-Hydrolyzing β-Lactamase Gene and of Two Novel Aminoglycoside Resistance Gene Cassettes
Authors: Poirel Laurent , Lambert Thierry , Türkoglü Salih , Ronco Esthel , Gaillard Jean-Louis , Nordmann Patrice 
Published: July 2002

ABSTRACT

Two clonally unrelated
Pseudomonas aeruginosa
clinical strains, RON-1 and RON-2, were isolated in 1997 and 1998 from patients hospitalized in a suburb of Paris, France. Both isolates expressed the class B carbapenem-hydrolyzing β-lactamase VIM-2 previously identified in Marseilles in the French Riviera. In both isolates, the
bla
VIM-2
cassette was part of a class 1 integron that also encoded aminoglycoside-modifying enzymes. In one case, two novel aminoglycoside resistance gene cassettes,
aacA29a
and
aacA29b
, were located at the 5′ and 3′ end of the
bla
VIM-2
gene cassette, respectively. The
aacA29a
and
aacA29b
gene cassettes were fused upstream with a 101-bp part of the 5′ end of the
qacE
cassette. The deduced amino acid sequence AAC(6′)-29a protein shared 96% identity with AAC(6′)-29b but only 34% identity with the
aacA7
-encoded AAC(6′)-I1, the closest relative of the AAC(6′)-I family enzymes. These aminoglycoside acetyltransferases had amino acid sequences much shorter (131 amino acids) than the other AAC(6′)-I enzymes (144 to 153 amino acids). They conferred resistance to amikacin, isepamicin, kanamycin, and tobramycin but not to gentamicin, netilmicin, and sisomicin.