bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effects of charged amino acids at b and c heptad positions on specificity and stability of four‐chain coiled coils
Authors: Vu Colynda , Robblee James , Werner Karin M. , Fairman Robert 
Published: July 2002

AbstractAn understanding of the balance of chemical forces responsible for protein stability and specificity of structure is essential for the success of efforts in protein design. Specifically, electrostatic interactions between charged amino acids have been explored extensively to understand the contribution of this force to protein stability. Much research on the importance of electrostatic interactions as specificity and stability determinants in two‐stranded coiled coils has been done, but there remains significant controversy about the magnitude of the attractive forces using such systems. We have developed a four‐stranded coiled‐coil system with charged residues incorporated at b and c heptad positions to explore the role of charge interactions. Here, we test quantitatively the effects of varying sidechain length on the magnitude of such electrostatic interactions. We synthesized peptides containing either aspartate or ornithine at both b and c heptad positions and tested their ability to self‐associate and to hetero‐associate with one another and with peptides containing glutamate or lysine at the same positions. We find that interactions between glutamate and either lysine or ornithine are more favorable than the corresponding interactions involving aspartate. In each case, charged interactions provide additional stability to coiled coils, although helix propensity effects may play a significant role in determining the overall stability of these structures.
Analysis of inhibitor binding in influenza virus neuraminidase
Authors: Smith Brian J. , Colman Peter M. , Von Itzstein Mark , Danylec Basil , Varghese Joseph N. 
Published: July 2002

Abstract2,3‐didehydro‐2‐deoxy‐N‐acetylneuraminic acid (DANA) is a transition state analog inhibitor of influenza virus neuraminidase (NA). Replacement of the hydroxyl at the C9 position in DANA and 4‐amino‐DANA with an amine group, with the intention of taking advantage of an increased electrostatic interaction with a conserved acidic group in the active site to improve inhibitor binding, significantly reduces the inhibitor activity of both compounds. The three‐dimensional X‐ray structure of the complexes of these ligands and NA was obtained to 1.4 Å resolution and showed that both ligands bind isosterically to DANA. Analysis of the geometry of the ammonium at the C4 position indicates that Glu119 may be neutral when these ligands bind. A computational analysis of the binding energies indicates that the substitution is successful in increasing the energy of interaction; however, the gains that are made are not sufficient to overcome the energy that is required to desolvate that part of the ligand that comes in contact with the protein.
Constrained modeling of spin–labeled major coat protein mutants from M13 bacteriophage in a phospholipid bilayer
Authors: Bashtovyy Denys , Marsh Derek , Hemminga Marcus A. , PÁLI Tibor 
Published: July 2002

AbstractThe family of three‐dimensional molecular structures of the major coat protein from the M13 bacteriophage, which was determined in detergent micelles by NMR methods, has been analyzed by constrained geometry optimization in a phospholipid environment. A single‐layer solvation shell of dioleoyl phosphatidylcholine lipids was built around the protein, after replacing single residues by cysteines with a covalently attached maleimide spin label. Both the residues substituted and the phospholipid were chosen for comparison with site‐directed spin labeling EPR measurements of distance and local mobility made previously on membranous assemblies of the M13 coat protein purified from viable mutants. The main criteria for identifying promising candidate structures, out of the 300 single‐residue mutant models generated for the membranous state, were 1) lack of steric conflicts with the phospholipid bilayer, 2) good match of the positions of spin‐labeled residues along the membrane normal with EPR measurements, and 3) a good match between the sequence profiles of local rotational freedom and a structural restriction parameter for the spin‐labeled residues obtained from the model. A single subclass of structure has been identified that best satisfies these criteria simultaneously. The model presented here is useful for the interpretation of future experimental data on membranous M13 coat protein systems. It is also a good starting point for full‐scale molecular dynamics simulations and for the design of further site‐specific spectroscopic experiments.
A Method to Accelerate Convergence and to Preserve Radiative Energy
Balance in Solving the P1 Equation by Iterative
Methods
Authors: Li Genong , Modest Michael F. 
Published: July 2002

Abstract
An acceleration method is proposed particularly for the P1 equation. The radiative energy balance is used as a constraint to correct iterative solutions. The method not only accelerates convergence but also preserves the radiative energy balance, the latter being of great importance when radiation calculations are coupled with flow calculations. This acceleration method can be applied to other elliptical problems with boundary conditions of the second and/or the third kind.
Midspan Flow-Field Measurements for Two Transonic Linear Turbine Cascades at Off-Design Conditions
Authors: Jouini D. B. M. , Sjolander S. A. , Moustapha S. H. 
Published: July 2002

Abstract
The paper presents detailed midspan experimental results from two transonic linear turbine cascades. The blades for the two cascades were designed for the same service and differ mainly in their leading-edge geometries. One of the goals of the study was to investigate the influence of the leading-edge metal angle on the sensitivity of the blade to positive off-design incidence. Measurements were made for incidence values of −10.0, 0.0, +4.5, +10.0, and +14.5 deg relative to design incidence. The exit Mach numbers varied roughly from 0.5 to 1.2 and the Reynolds numbers from about 4×105 to 106. The measurements include the midspan losses, blade loadings and base pressures. In addition, the axial-velocity-density ratio (AVDR) was extracted for each operating point. The AVDR was found to vary from about 0.98 at −10.0 deg of incidence to about 1.27 at +14.5 deg. Thus, the data set also provides some evidence of the influence AVDR on axial turbine blade performance. Detailed experimental results for turbine blade performance at off-design incidence are very scarce in the open literature, particularly for transonic conditions. Among other things, the present results are intended to expand the database available in the open literature. To this end, the key aerodynamic results are presented in tabular form, along with the detailed geometry of the cascades. The results could be used in the development of new or improved correlations for use in the early stages of design. They could also be used to evaluate the ability of current CFD codes to capture reliably the variation in losses and other aerodynamic quantities with variations in blade incidence.
Biological Cost of AmpC Production for
Salmonella enterica
Serotype Typhimurium
Authors: Morosini M. I. , Ayala J. A. , Baquero F. , Martínez J. L. , Blázquez J. 
Published: July 2002

ABSTRACT

Chromosomally mediated AmpC-type β-lactamases are frequently found among
Enterobacteriaceae
. Hyperproduction of AmpC β-lactamase results in high-level resistance to β-lactam antibiotics. One striking feature of
Salmonella
is the absence of the structural
ampC
gene, encoding AmpC β-lactamase, in contrast with other members in the
Enterobacteriaceae
family, such as
Escherichia
,
Citrobacter
, or
Enterobacter
. The horizontal acquisition of
ampC
genes is one of the causes of the increased resistance to extended-spectrum cephalosporins and β-lactamase inhibitors among gram-negative rods. Nevertheless, despite the high number of β-lactam-resistant
Salmonella
isolates so far described, only two strains expressing resistance to cephalosporin and β-lactamase inhibitors which is mediated by AmpC-type enzymes have been found. In this work, data are provided which support the possibility that the maintenance and expression of the
ampC
gene may represent an unbearable cost for
Salmonella
in terms of reduction of some of its lifestyle attributes, such as growth rate and invasiveness. The deleterious AmpC burden can be eliminated by decreasing the production of AmpC when both the regulatory gene,
ampR
, and
ampC
are present in
Salmonella
. Thus, it is suggested that the two genes have to be acquired together by
Salmonella
, leading to an inducible β-lactam resistance phenotype. AmpC synthesis did not produce major variations in the peptidoglycan composition of
Salmonella
.
Resistance to Methotrexate Due to AcrAB-Dependent Export from
Escherichia coli
Authors: Kopytek Stephan J. , Dyer John C. D. , Knapp Gwendowlyn S. , Hu James C. 
Published: July 2002

ABSTRACT

Many laboratory strains of
Escherichia coli
are resistant to methotrexate (MTX), a folate analogue that binds dihydrofolate reductase (DHFR). Mutations that inactivate either
tolC
or
acrA
confer MTX sensitivity. Further, overexpression of a fusion protein with DHFR activity reverses this sensitivity by titrating out intracellular MTX. These results suggest that MTX accumulates in cells where mutations in
acrA
or
tolC
have inactivated the TolC-dependent AcrAB multidrug resistance efflux pump.
A New SHV-Derived Extended-Spectrum β-Lactamase (SHV-24) That Hydrolyzes Ceftazidime through a Single-Amino-Acid Substitution (D179G) in the Ω-Loop
Authors: Kurokawa Hiroshi , Yagi Tetsuya , Shibata Naohiro , Shibayama Keigo , Kamachi Kazunari , Arakawa Yoshichika 
Published: July 2002

ABSTRACT

A new SHV-derived extended-spectrum β-lactamase (SHV-24) conferring high-level resistance to ceftazidime but not cefotaxime and cefazolin was identified in Japan. This enzyme was encoded by a transferable 150-kb plasmid from an
Escherichia coli
clinical isolate. The pI and

K
m

for CAZ of this enzyme were 7.5 and 30 μM, respectively. SHV-24 was found to have a D179G substitution in the Ω-loop of the enzyme.
Contribution of the MexX-MexY-OprM Efflux System to Intrinsic Resistance in
Pseudomonas aeruginosa
Authors: Masuda Nobuhisa , Sakagawa Eiko , Ohya Satoshi , Gotoh Naomasa , Tsujimoto Hideto , Nishino Takeshi 
Published: July 2002

ABSTRACT

To test the possibility that MexX-MexY, a new set of efflux system components, is associated with OprM and contributes to intrinsic resistance in
Pseudomonas aeruginosa
, we constructed a series of isogenic mutants lacking
mexXY
and/or
mexAB
and/or
oprM
from a laboratory strain PAO1, and examined their susceptibilities to ofloxacin, tetracycline, erythromycin, gentamicin, and streptomycin. Loss of either MexXY or OprM from the MexAB-deficient mutant increased susceptibility to all agents tested, whereas loss of MexXY from the MexAB-OprM-deficient mutant caused no change in susceptibility. Introduction of an OprM expression plasmid decreased the susceptibility of the
mexAB-oprM
-deficient-/
mexXY
-maintaining mutant, yet caused no change in the susceptibility of a
mexAB-oprM
- and
mexXY
-deficient double mutant. Immunoblot analysis using anti-MexX polyclonal rabbit serum generated against synthetic oligopeptides detected expression of MexX in the PAO1 cells grown in medium containing tetracycline, erythromycin, or gentamicin, although expression of MexX was undetectable in the cells incubated in medium without any agent. These results suggest that MexXY induced by these agents is functionally associated with spontaneously expressed OprM and contributes to the intrinsic resistance to these agents.
Concentration-Dependent Selection of Small Phenotypic Differences in TEM β-Lactamase-Mediated Antibiotic Resistance
Authors: Negri Maria-Cristina , Lipsitch Marc , Blázquez Jesús , Levin Bruce R. , Baquero Fernando 
Published: July 2002

ABSTRACT

In this paper, the first robust experimental evidence of in vitro and in vivo concentration-dependent selection of low-level antibiotic-resistant genetic variants is described. The work is based on the study of an asymmetric competition assay with pairs of isogenic
Escherichia coli
strains, differing only (apart from a neutral chromosomal marker) in a single amino acid replacement in a plasmid-mediated TEM-1 beta-lactamase enzyme, which results in the new TEM-12 beta-lactamase. The mixture was challenged by different antibiotic concentrations, both in vitro and in the animal model, and the selective process of the variant population was carefully monitored. A mathematical model was constructed to test the hypothesis that measured growth and killing rates of the individual TEM variants at different antibiotic concentrations could be used to predict quantitatively the strength of selection for TEM-12 observed in competition experiments at these different concentrations.
A Ribosomal ATPase Is a Target for Hygromycin B Inhibition on
Escherichia coli
Ribosomes
Authors: Ganoza M. Clelia , Kiel Michael C. 
Published: July 2002

ABSTRACT

We demonstrate that the transfer of fully charged aminoacyl-tRNAs into peptides directed by the MS2 RNA template requires both ATP and GTP, initiation factors (IF1, IF2, and IF3), elongation factors (EF-Tu, EF-Ts, and EF-G), and the ribosomal ATPase (RbbA). The nonhydrolyzable analogue AMPPCP inhibits the reactions, suggesting that hydrolysis of ATP is required for synthesis. The RbbA protein occurs bound to ribosomes and stimulates the ATPase activity of
Escherichia coli
70S and 30S particles. The gene encoding RbbA harbors four ATP binding domains; the C-terminal half of the protein bears extensive sequence similarity to EF-3, a ribosome-dependent ATPase. Here, we show that the antibiotic hygromycin B selectively inhibits the ATPase activity of RbbA. Other antibiotics with similar effects on miscoding, streptomycin and neomycin, as well as antibiotics that impair peptide bond synthesis and translocation, had little effect on the ATPase activity of RbbA on 70S ribosomes. Immunoblot analysis indicates that at physiological concentrations, hygromycin B selectively releases RbbA from 70S ribosomes. Hygromycin B protects G1494 and A1408 in the decoding region, and RbbA enhances the reactivity of A889 and G890 of the 16S rRNA switch helix region. Cross-linking and X-ray diffraction data have revealed that this helix switch and the decoding region are in close proximity. Mutations in the switch helix (889-890) region affect translational fidelity and translocation. The binding site of hygromycin B and its known dual effect on the fidelity of decoding and translocation suggest a model for the action of this drug on ribosomes.
Rifampin Resistance in
Mycobacterium kansasii
Is Associated with
rpoB
Mutations
Authors: Klein John L. , Brown Timothy J. , French Gary L. 
Published: July 2002

ABSTRACT

Rifampin is the most potent drug used in the treatment of disease due to
Mycobacterium kansasii
. A 69-bp fragment of
rpoB
, the gene that encodes the β subunit of the bacterial RNA polymerase, was sequenced and found to be identical in five rifampin-susceptible clinical isolates of
M. kansasii
. This sequence showed 87% homology with the
Mycobacterium tuberculosis
gene, with an identical deduced amino acid sequence. In contrast, missense mutations were detected in the same fragment amplified from five rifampin-resistant isolates. A rifampin-resistant strain generated in vitro also harbored an
rpoB
gene missense mutation that was not present in the parent isolate. All mutations detected (in codons 513, 526, and 531) have previously been described in rifampin-resistant
M. tuberculosis
isolates. Rifampin MICs determined by E-test were <1 mg/liter for all rifampin-susceptible isolates and >256 mg/liter for all rifampin-resistant ones. In addition, four of the five rifampin-resistant isolates were also resistant to rifabutin. We have thus shown a strong association between
rpoB
gene missense mutations and rifampin resistance in
M. kansasii
. Although our results are derived from a small number of isolates and confirmation with larger numbers would be useful, they strongly suggest that mutations within
rpoB
form the molecular basis of rifampin resistance in this species.
CTX-M-Type Extended-Spectrum β-Lactamase That Hydrolyzes Ceftazidime through a Single Amino Acid Substitution in the Omega Loop
Authors: Poirel Laurent , Naas Thierry , Le Thomas Isabelle , Karim Amal , Bingen Edouard , Nordmann Patrice 
Published: July 2002

ABSTRACT

Escherichia coli
ILT-1,
Klebsiella pneumoniae
ILT-2, and
K. pneumoniae
ILT-3 were isolated in May 1999 in Paris, France, from a rectal swab of a hospitalized 5-month-old girl. These isolates had a clavulanic acid-inhibited substrate profile that included expanded-spectrum cephalosporins. The MICs of cefotaxime were higher for
E. coli
ILT-1 and
K. pneumoniae
ILT-2 than for
K. pneumoniae
ILT-3, while the opposite was found for the MICs of ceftazidime. Genetic and biochemical analyses revealed that
E. coli
ILT-1 and
K. pneumoniae
ILT-2 produced the CTX-M-18 β-lactamase, while
K. pneumoniae
ILT-3 produced the CTX-M-19 β-lactamase. The amino acid sequence of the CTX-M-18 β-lactamase differed from that of the CTX-M-9 β-lactamase by an Ala-to-Val change at position 231, while CTX-M-19 possessed an additional Pro-to-Ser change at position 167 in the omega loop of Ambler class A enzymes. The latter amino acid substitution may explain the CTX-M-19-mediated hydrolysis of ceftazidime, which has not been reported for other CTX-M-type enzymes. The
bla
CTX-M-18
and
bla
CTX-M-19
genes were located on transferable plasmids that varied in size (ca. 60 and 50 kb, respectively) but that showed similar restriction patterns.
Resistance-Nodulation-Cell Division-Type Efflux Pump Involved in Aminoglycoside Resistance in
Acinetobacter baumannii
Strain BM4454
Authors: Magnet Sophie , Courvalin Patrice , Lambert Thierry 
Published: July 2002

ABSTRACT

Multidrug-resistant strain
Acinetobacter baumannii
BM4454 was isolated from a patient with a urinary tract infection. The
adeB
gene, which encodes a resistance-nodulation-cell division (RND) protein, was detected in this strain by PCR with two degenerate oligodeoxynucleotides. Insertional inactivation of
adeB
in BM4454, which generated BM4454-1, showed that the corresponding protein was responsible for aminoglycoside resistance and was involved in the level of susceptibility to other drugs including fluoroquinolones, tetracyclines, chloramphenicol, erythromycin, trimethoprim, and ethidium bromide. Study of ethidium bromide accumulation in BM4454 and BM4454-1, in the presence or in the absence of carbonyl cyanide
m
-chlorophenylhydrazone, demonstrated that AdeB was responsible for the decrease in intracellular ethidium bromide levels in a proton motive force-dependent manner. The
adeB
gene was part of a cluster that included
adeA
and
adeC
which encodes proteins homologous to membrane fusion and outer membrane proteins of RND-type three-component efflux systems, respectively. The products of two upstream open reading frames encoding a putative two-component regulatory system might be involved in the regulation of expression of the
adeABC
gene cluster.
Thermoreversible Gel Formulations Containing Sodium Lauryl Sulfate or
n
-Lauroylsarcosine as Potential Topical Microbicides against Sexually Transmitted Diseases
Authors: Roy Sylvie , Gourde Pierrette , Piret Jocelyne , Désormeaux André , Lamontagne Julie , Haineault Caroline , Omar Rabeea F. , Bergeron Michel G. 
Published: July 2002

ABSTRACT

The microbicidal efficacies of two anionic surfactants, sodium lauryl sulfate (SLS) and
n
-lauroylsarcosine (LS), were evaluated in cultured cells and in a murine model of herpes simplex type 2 (HSV-2) intravaginal infection. In vitro studies showed that SLS and LS were potent inhibitors of the infectivity of HSV-2 strain 333. The concentrations of SLS which inhibit viral infectivity by 50% (50% inhibitory dose) and 90% (90% inhibitory dose) were 32.67 and 46.53 μM, respectively, whereas the corresponding values for LS were 141.76 and 225.30 μM. In addition, intravaginal pretreatment of mice with thermoreversible gel formulations containing 2.5% SLS or 2.5% LS prior to the inoculation of HSV-2 strain 333 completely prevented the development of genital herpetic lesions and the lethality associated with infection. Of prime interest, no infectious virus could be detected in mouse vaginal mucosa. Both formulations still provided significant protection when viral challenge was delayed until 1 h after pretreatment. Finally, intravaginal application of gel formulations containing 2.5% SLS or 2.5% LS once daily for 14 days to rabbits did not induce significant irritations to the genital mucosa, as demonstrated from macroscopic and histopathologic examinations. These results suggest that thermoreversible gel formulations containing SLS or LS could represent potent and safe topical microbicides for the prevention of HSV-2 and possibly other sexually transmitted pathogens, including human immunodeficiency virus.
Desbutyl-Benflumetol, a Novel Antimalarial Compound: In Vitro Activity in Fresh Isolates of
Plasmodium falciparum
from Thailand
Authors: Noedl Harald , Allmendinger Thomas , Prajakwong Somsak , Wernsdorfer Gunther , Wernsdorfer Walther H. 
Published: July 2002

ABSTRACT

Desbutyl-benflumetol (DBB) is a novel antimalarial compound closely related to benflumetol (lumefantrine), of which it is a putative metabolite. The in vitro response of
Plasmodium falciparum
to DBB was studied in Mae Hong Son and Mae Sot, in northwest Thailand, in 1997 and 1998. In total, 155 fresh isolates were successfully tested using the World Health Organization standard in vitro microtest system (Mark II). The mean 50% effective concentration (EC
50
) and 90% effective concentration of DBB were 6.36 and 31.09 nmol/liter, respectively. The comparison of the activity of DBB and benflumetol yielded a highly significant potency ratio of 4.52, corresponding to a more than four times higher efficacy of DBB. A considerable potency difference was found between isolates from Mae Hong Son and those from Mae Sot, reflecting lesser sensitivity in the area with marked resistance to mefloquine and quinine. This observation is also supported by a highly significant activity correlation with benflumetol (
P
< 0.001) and to a similar degree with mefloquine (
P
< 0.001), reflecting a close relationship of DBB with the class II aryl amino alcohol blood schizontocides. A less distinct association was also found with artemisinin, which was significant only at the EC
50
level, and there was no correlation at all with chloroquine. DBB is a promising antimalarial compound that merits further investigation in order to define its practical therapeutic potential.