bluuebunny/crossref_metadata_2025_split huggingface dataset:
Sequencing of the ddl gene and modeling of the mutated D‐alanine:D‐alanine ligase in glycopeptide‐dependent strains of Enterococcus faecium
Authors: Gholizadeh Y. , Prevost M. , Van Bambeke F. , Casadewall B. , Tulkens P.M. , Courvalin P. 
Published: July 2002

AbstractGlycopeptide dependence for growth in enterococci results from mutations in the ddl gene that inactivate the host D‐Ala:D‐Ala ligase. The strains require glycopeptides as inducers for synthesis of resistance proteins, which allows for the production of peptidoglycan precursors ending in D‐Ala‐D‐Lac instead of D‐Ala‐D‐Ala. The sequences of the ddl gene from nine glycopeptide‐dependent Enterococcus faecium clinical isolates were determined. Each one had a mutation consisting either in a 5‐bp insertion at position 41 leading to an early stop codon, an in‐frame 6‐bp deletion causing the loss of two residues (KDVA243‐246 to KA), or single base‐pair changes resulting in an amino acid substitution (E13 → G, G99 → R, V241 → D, D295 → G, P313 → L). The potential consequences of the deletion and point mutations on the 3‐D structure of the enzyme were evaluated by comparative molecular modeling of the E. faecium enzyme, using the X‐ray structure of the homologous Escherichia coli D‐Ala:D‐Ala ligase DdlB as a template. All mutated residues were found either to interact directly with one of the substrates of the enzymatic reaction (E13 and D295) or to stabilize the position of critical residues in the active site. Maintenance of the 3‐D structure in the vicinity of these mutations in the active site appears critical for D‐Ala:D‐Ala ligase activity.
Conformational characterization of oligomeric intermediates and aggregates in β‐lactoglobulin heat aggregation
Authors: Carrotta Rita , Bauer Rogert , Waninge Rianne , Rischel Christian 
Published: July 2002

AbstractIn one of the first studies of isolated intermediates in protein aggregation, we have used circular dichroism and fluorescence spectroscopy to characterize metastable oligomers that are formed in the early steps of β‐lactoglobulin heat aggregation. The intermediates show typical molten globule characteristics (secondary structure content similar to the native and less tight packing of the side chains), in agreement with the belief that partly folded states play a key role in protein aggregation. The far‐UV CD signal bears strong resemblance to that of a known folding intermediate. Cryo‐transmission electron microscopy of the aggregates reveals spherical particles with a diameter of about 50 nm and an internal threadlike structure. Isolated oligomers as well as larger aggregates bind the dye thioflavin T, usually a signature of the amyloid superstructures found in many protein aggregates. This result suggests that the structural motif recognized by thioflavin T can be formed in small oligomers.
Thymidylate kinase of Mycobacterium tuberculosis: A chimera sharing properties common to eukaryotic and bacterial enzymes
Authors: Munier‐Lehmann HÉLÈNE , Chaffotte Alain , Pochet Sylvie , Labesse Gilles 
Published: July 2002

AbstractWe have overexpressed in Escherichia coli the thymidylate kinase of Mycobacterium tuberculosis (TMPKmt). Biochemical and physico‐chemical characterization of TMPKmt revealed distinct structural and catalytic features when compared to its counterpart from yeast (TMPKy) or E. coli (TMPKec). Denaturation of the dimeric TMPKmt by urea under equilibrium conditions was studied by intrinsic fluorescence and circular dichroism (CD) spectroscopy. It suggested a three‐state unfolding mechanism with a monomeric intermediate. On the other hand, 3′‐azido‐3′‐deoxythymidine monophosphate (AZT‐MP), which is substrate for TMPKy and TMPKec acts as a potent competitive inhibitor for TMPKmt. We propose a structural model of TMPKmt in which the overall fold described in TMPKy and TMPKec is conserved and slight differences at the level of primary and 3D‐structure explain strong variations in the phosphorylation rate of substrate analogs. According to the model, we synthesized dTMP analogs acting either as substrates or specific inhibitors of TMPKmt. This approach based on slight structural differences among similar proteins could be applied to other essential enzymes for the design of new species‐specific antimicrobials.
Photoacid bulkiness effect on dissolution kinetics in chemically amplified deep ultraviolet resists
Authors: Itani Toshiro , Yoshino Hiroshi , Fujimoto Masashi , Kasama Kunihiko 
Published: July 2002

The photoacid bulkiness effect on dissolution characteristics was investigated in a three-component chemically amplified deep ultraviolet positive resist system. The resist consisted of a tert-butoxycarbonyl (t-BOC) protected polystyrene, benzenesulfonic acid derivative photoacid generator and an additional dissolution inhibitor (t-BOC protected bisphenol A). For the benzenesulfonic acid photoacid generator, four types of substituents; 4-fluoro, 4-chloro, 2,4-dimethyl, and 4-tert-butyl, were studied. Dissolution contrast (Rmax/Rmin) increased and the pattern profiles became rectangular with increasing substituent bulkiness. The reason for this is that bulky substituents such as 2,4-dimethyl and 4-tert-butyl increased resist hydrophobic strength in both bulk and surface regions, thereby strengthening the surface inhibition effect. Next, in order to investigate the bulkiness effect on acid mobility, diffusion length during postexposure bake time was determined by means of the ion conductivity method. Diffusion length decreased and resist profiles became rectangular with increasing photoacid bulkiness, and T-topping profile was observed in 4-tert-butyl substituent. It is believed that the acid catalytic reaction number decreases in the case of a shorter diffusion length. Therefore, the surface inhibition effect becomes strong, and then the T-topping profile was observed in 4-tert-butyl substituent.
In order to understand the details of surface inhibition effect, resist pattern profile simulation was carried out using PROLITH/2 version 3.0. Simulation results of pattern profiles coincided with the actual resist pattern by adjusting the parameters of the surface inhibition effect (relative surface rate and inhibition depth). It was found that the photoacid bulkiness effect on pattern profiles can be explained by the surface inhibition effect. Based on the analysis of the dissolution characteristics, acid diffusion behavior and profile simulation, it was concluded that the resist pattern profile, especially its top rectangularity, can be controlled by changing the substituent group of photogenerated benzenesulfonic acid. By combining the modification of the base resin structure (protection ratio and molecular weight) and acid structures, both high resolution capability and rectangular resist pattern profiles can be achieved.
Antimycobacterial Activities of 2,4-Diamino-5-Deazapteridine Derivatives and Effects on Mycobacterial Dihydrofolate Reductase
Authors: Suling William J. , Seitz Lainne E. , Pathak Vibha , Westbrook Louise , Barrow Esther W. , Zywno-van-Ginkel Sabrina , Reynolds Robert C. , Piper J. Robert , Barrow William W. 
Published: July 2002

ABSTRACT

Development of new antimycobacterial agents for
Mycobacterium avium
complex (MAC) infections is important particularly for persons coinfected with human immunodeficiency virus. The objectives of this study were to evaluate the in vitro activity of 2,4-diamino-5-methyl-5-deazapteridines (DMDPs) against MAC and to assess their activities against MAC dihydrofolate reductase recombinant enzyme (rDHFR). Seventy-seven DMDP derivatives were evaluated initially for in vitro activity against one to three strains of MAC (NJ168, NJ211, and/or NJ3404). MICs were determined with 10-fold dilutions of drug and a colorimetric (Alamar Blue) microdilution broth assay. MAC rDHFR 50% inhibitory concentrations versus those of human rDHFR were also determined. Substitutions at position 5 of the pteridine moiety included -CH
3
, -CH
2
CH
3
, and -CH
2
OCH
3
groups. Additionally, different substituted and unsubstituted aryl groups were linked at position 6 through a two-atom bridge of either -CH
2
NH, -CH
2
N(CH
3
), -CH
2
CH
2
, or -CH
2
S. All but 4 of the 77 derivatives were active against MAC NJ168 at concentrations of ≤13 μg/ml. Depending on the MAC strain used, 81 to 87% had MICs of ≤1.3 μg/ml. Twenty-one derivatives were >100-fold more active against MAC rDHFR than against human rDHFR. In general, selectivity was dependent on the composition of the two-atom bridge at position 6 and the attached aryl group with substitutions at the 2′ and 5′ positions on the phenyl ring. Using this assessment, a rational synthetic approach was implemented that resulted in a DMDP derivative that had significant intracellular activity against a MAC-infected Mono Mac 6 monocytic cell line. These results demonstrate that it is possible to synthesize pteridine derivatives that have selective activity against MAC.
Detection of the High-Level Aminoglycoside Resistance Gene
aph(2")-Ib
in
Enterococcus faecium
Authors: Kao Susan J. , You Il , Clewell Don B. , Donabedian Susan M. , Zervos Marcus J. , Petrin Joanne , Shaw Karen J. , Chow Joseph W. 
Published: July 2002

ABSTRACT

A new high-level gentamicin resistance gene, designated
aph(2")-Ib
, was cloned from
Enterococcus faecium
SF11770. The deduced amino acid sequence of the 897-bp open reading frame of
aph(2")-Ib
shares homology with the aminoglycoside-modifying enzymes AAC(6′)-APH(2"), APH(2")-Ic, and APH(2")-Id. The observed phosphotransferase activity is designated APH(2")-Ib.
Multiple-Antibiotic Resistance Mediated by Structurally Related IncL/M Plasmids Carrying an Extended-Spectrum β-Lactamase Gene and a Class 1 Integron
Authors: Villa Laura , Pezzella Cristina , Tosini Fabio , Visca Paolo , Petrucca Andrea , Carattoli Alessandra 
Published: July 2002

ABSTRACT

A conjugative IncL/M plasmid (pSEM) conferring resistance to gentamicin, amikacin, kanamycin, sulfonamides, and expanded-spectrum cephalosporins was found in pathogenic strains of
Salmonella enterica
serotype Typhimurium. Resistance to aminoglycosides was encoded by a
sul1
-type class 1 integron (In-t3). An extended-spectrum beta-lactamase gene,
bla
SHV-5
, was identified 3.5 kb downstream of the integrase (
intI1
) gene of In-t3. Nucleotide sequence analysis of the 5.3-kb
bla
SHV-5
–In-t3 region of pSEM highlighted striking similarities with IncL/M plasmids isolated from nosocomial gram-negative pathogens, conferring resistance to expanded-spectrum cephalosporins and aminoglycosides.
Activity of LY333328 Combined with Gentamicin In Vitro and in Rabbit Experimental Endocarditis Due to Vancomycin-Susceptible or -Resistant
Enterococcus faecalis
Authors: Lefort Agnès , Saleh-Mghir Azzam , Garry Louis , Carbon Claude , Fantin Bruno 
Published: July 2002

ABSTRACT

We investigated the activity of LY333328 alone and combined with gentamicin, both in vitro and in a rabbit model of experimental endocarditis, against the susceptible strain
Enterococcus faecalis
JH2-2 and its two glycopeptide-resistant transconjugants, BM4316 (VanA) and BM4275 (VanB). MICs of LY333328 and gentamicin were 2 and 16 μg/ml, respectively, for the three strains. In vitro, LY333328 alone was bactericidal at 24 h against JH2-2 at a concentration of 2 μg/ml and against BM4316 and BM4275 at a concentration of 30 μg/ml. The combination of LY333328 and gentamicin (4 μg/ml) was synergistic and bactericidal after 24 h of incubation against the three strains at LY333328 concentrations of 2 μg/ml for JH2-2 and 8 μg/ml for BM4275 and BM4316. The combination of LY333328 and gentamicin was the only regimen demonstrating in vitro bactericidal activity against BM4316. In vivo, intravenous treatment with LY333328 alone, providing peak and trough serum levels of 83.3 ± 1.3 and 3.8 ± 0.2 μg/ml, respectively, was inactive against BM4316 and BM4275 and selected mutants resistant to LY333328 in half of the rabbits infected with the VanA-type strain (MICs, 8 to 20 μg/ml). However, the LY333328-gentamicin combination was active against the three strains and prevented the emergence of mutants resistant to both components of the combination. We conclude that the LY333328-gentamicin combination might be of interest for the treatment of enterococcal infections, particularly against VanA-type strains.
Therapeutic Efficacy of GAR-936, a Novel Glycylcycline, in a Rat Model of Experimental Endocarditis
Authors: Murphy Timothy M. , Deitz Jacqueline M. , Petersen Peter J. , Mikels Susan M. , Weiss William J. 
Published: July 2002

ABSTRACT

GAR-936, a novel glycylcycline, was investigated with a rat model of experimental endocarditis. It was compared with vancomycin against both vancomycin-susceptible and -resistant
Enterococcus faecalis
and methicillin-resistant
Staphylococcus aureus
. GAR-936 exhibited the lowest MICs (≤0.12 μg/ml) in vitro against each of the isolates tested. Endocarditis was established by placement of a catheter across the aortic valve, followed by intravenous injection of 10
6
CFU of bacteria 48 h later. Treatment with GAR-936 or vancomycin was initiated 24 to 36 h after bacterial infection and administered subcutaneously twice a day for 3 days at ascending doses. GAR-936 reduced bacterial vegetation titers by >2 log
10
CFU, compared to those in untreated controls, for both vancomycin-susceptible and -resistant (VanA and VanB)
E. faecalis
strains and >4 log
10
CFU for a methicillin-resistant
S. aureus
isolate. The glycylcycline was more efficacious at a lower administered dose in the rat model of endocarditis than was vancomycin. The efficacy of GAR-936 in this model was apparently not enhanced by a factor in rat serum, as was observed for vancomycin with a time-kill curve. The results of this study demonstrate the therapeutic potential of GAR-936 for the treatment of enterococcal and staphylococcal infections and warrant further investigation.
Low Level of Cross-Resistance to Amprenavir (141W94) in Samples from Patients Pretreated with Other Protease Inhibitors
Authors: Schmidt Barbara , Korn Klaus , Moschik Brigitte , Paatz Christiane , Überla Klaus , Walter Hauke 
Published: July 2002

ABSTRACT

The therapeutic success of an antiretroviral salvage regimen containing protease inhibitors (PI) is limited by PI-resistant viral strains exhibiting various degrees of resistance and cross-resistance. To evaluate the extent of cross-resistance to the new PI amprenavir, 155 samples from 132 human immunodeficiency virus type 1-infected patients were analyzed for viral genotype by direct sequencing of the protease gene. Concomitantly, drug sensitivity to indinavir, saquinavir, ritonavir, nelfinavir, and amprenavir was analyzed by a recombinant virus assay. A total of 111 patients had been pretreated with 1-4 PI, but all were naive to amprenavir. A total of 105 samples (67.7%) were sensitive to amprenavir; 25 samples (16.1%) were intermediately resistant, and another 25 samples were highly resistant (4- to 8-fold- and >8-fold-reduced sensitivity, respectively). The mutations 46I/L, 54L/V, 84V, and 90M showed the strongest association with amprenavir resistance (
P
< 0.0001). The scoring system using 84V and/or any two of a number of mutations (10I/R/V/F, 46I/L, 54L/V, and 90M) predicted amprenavir resistance with a sensitivity of 86.0% and a specificity of 81.0% within the analyzed group of samples. Of 62 samples with resistance against 4 PI, 23 (37.1%) were still sensitive to amprenavir. In comparison, only 2 of 23 samples (8.7%) from nelfinavir-naive patients with resistance against indinavir, saquinavir, and ritonavir were still sensitive to nelfinavir. Amprenavir thus appears to be an interesting alternative for PI salvage therapy.
Characterization of Novel Human Hepatoma Cell Lines with Stable Hepatitis B Virus Secretion for Evaluating New Compounds against Lamivudine- and Penciclovir-Resistant Virus
Authors: Fu Lei , Cheng Yung-Chi 
Published: July 2002

ABSTRACTl-Nucleoside analogs are new therapeutic agents for treatment of chronic hepatitis B. However, their clinical application was limited by the emergence of viral resistance. It is important to develop a new system to evaluate drug cross-resistance and to test new agents that may overcome resistant virus. In this report, three cell lines HepG2-WT10, HepG2-SM1, and HepG2-DM2 are presented; these cell lines were established by transfection of HepG2 cells with unique fully functional 1.1× hepatitis B virus (HBV) genomes: wild-type HBV-adr and its L526M and L526MM550V variants, respectively. We have demonstrated that these genomes have different susceptibilities to lamivudine [l(−)SddC] and penciclovir (PCV). By examining HBV RNA transcription, antigen expression, progeny DNA replication, and viral susceptibilities tol(−)SddC, PCV, and other nucleoside analogs, it is concluded that the cell lines are able to stably producel(−)SddC- and PCV-sensitive and -resistant HBV virions. In addition, the relative susceptibilities of the wild-type and mutant HBV produced from the stably transfected cell lines to several anti-HBV nucleoside analogs were also examined and found to be about the same as those found by using a transient infection system. PMEA [9-(2-phosphonylmethoxytehyl)-adenine] and QYL685 are able to suppressl(−)SddC- and PCV-resistant HBV. In conclusion, this cell culture system is a novel and useful tool for evaluating anti-HBV compounds and biologics.
Molecular Mechanisms of Fluoroquinolone Resistance in
Pseudomonas aeruginosa
Isolates from Cystic Fibrosis Patients
Authors: Jalal Shah , Ciofu Oana , Høiby Niels , Gotoh Naomasa , Wretlind Bengt 
Published: July 2002

ABSTRACT

Twenty
P. aeruginosa
isolates were collected from six cystic fibrosis (CF) patients, aged 27 to 33, in 1994 (9 isolates) and 1997 (11 isolates) at the CF Center, Copenhagen, Denmark, and were typed by pulse-field gel electrophoresis (PFGE) or ribotyping. Five of the patients had isolates with the same PFGE or ribotyping patterns in 1997 as in 1994, and ciprofloxacin had a two- to fourfold higher MIC for the isolates collected in 1997 than those from 1994. Genomic DNA was amplified for
gyrA
,
parC
,
mexR
, and
nfxB
by PCR and sequenced. Eleven isolates had mutations in
gyrA
, seven isolates had mutations at codon 83 (Thr to Ile), and four isolates had mutations at codon 87 (Asp to Asn or Tyr). Sixteen isolates had mutations in
nfxB
at codon 82 (Arg to Leu). Increased amounts of OprN were found in six isolates and OprJ in eight isolates as determined by immunoblotting. No isolates had mutations in
parC
or
mexR
. Six isolates had mutations in efflux pumps without
gyrA
mutations. The average number of mutations was higher in isolates from 1997 than in those from 1994. The results also suggested that efflux resistance mechanisms are more common in isolates from CF patients than in strains from urine and wounds from non-CF patients, in which mutations in
gyrA
and
parC
dominate (S. Jalal and B. Wretlind, Microb. Drug Resist. 4:257–261, 1998).
Antiviral Activity of 2′-Deoxy-3′-Oxa-4′-Thiocytidine (BCH-10652) against Lamivudine-Resistant Human Immunodeficiency Virus Type 1 in SCID-hu Thy/Liv Mice
Authors: Stoddart Cheryl A. , Moreno Mary E. , Linquist-Stepps Valerie D. , Bare Christopher , Bogan Mark R. , Gobbi Alberto , Buckheit Robert W. , Bedard Jean , Rando Robert F. , McCune Joseph M. 
Published: July 2002

ABSTRACT
Oral administration of 2′-deoxy-3′-oxa-4′-thiocytidine (BCH-10652), a nucleoside analog structurally similar to lamivudine (3TC), caused dose-dependent inhibition of viral replication in SCID-hu Thy/Liv mice infected with human immunodeficiency virus type 1 NL4-3 and with an NL4-3 clone containing the M184V mutation in reverse transcriptase that confers resistance to 3TC. These experiments demonstrate the utility of this mouse model for evaluating drug resistance and for performing direct comparisons between antiviral compounds in vivo.
Identification of a Streptogramin A Acetyltransferase Gene in the Chromosome of
Yersinia enterocolitica
Authors: Seoane Asunción , García Lobo Juan M. 
Published: July 2002

ABSTRACT

Streptogramins are polypeptide antibiotics inhibiting protein synthesis by the prokaryotic ribosome. Gram-positive organisms are susceptible to streptogramins, while most gram-negative bacteria are intrinsically resistant. We have found a genomic fragment from a
Yersinia enterocolitica
isolate with an open reading frame coding for a polypeptide similar to the virginiamycin acetyltransferases found in various plasmids from gram-positive bacteria. The susceptible
Escherichia coli
strain DB10 was transformed to resistance to the type A streptogramins and to mixed (A + B) streptogramins upon introduction of a plasmid containing that gene. In addition, we showed streptogramin acetylating activity in vitro dependent on the presence of the
Y. enterocolitica sat
gene. Southern blot hybridization experiments showed that the
sat
gene was present in all the
Y. enterocolitica
isolates examined. These data together show that the gene in the
Y. enterocolitica
chromosome encoded an active streptogramin acetyltransferase. The deduced sequence of the
Y. enterocolitica
Sat protein was close to those of
sat
gene products found in gram-positive bacteria and cyanobacteria, suggesting a common evolutionary origin.
Pharmacodynamics of Glycopeptides in the Mouse Peritonitis Model ofStreptococcus pneumoniaeorStaphylococcus aureusInfection
Authors: Knudsen Jenny Dahl , Fuursted Kurt , Raber Susan , Espersen Frank , Frimodt-Møller Niels 
Published: July 2002

ABSTRACTThe emergence of resistance to various antibiotics in pneumococci leaves the glycopeptides as the only antibiotics against which pneumococci have no resistance mechanism. This situation has led to a renewed interest in the use of glycopeptides. It has not yet been possible to conclude which one or more of the pharmacokinetic or pharmacodynamic (PK/PD) parameters are the most important and best predictors for the effects of treatment with glycopeptides in animal models or in humans. We used the mouse peritonitis model with immunocompetent mice and withStaphylococcus aureusandStreptococcus pneumoniaeas infective organisms. A wide spectrum of different treatment regimens with vancomycin and teicoplanin was tested to study the pharmacodynamics of these drugs. In studies in which the single dose that protected 50% of lethally infected mice (ED50) was given as one dose or was divided into two doses, survival was significantly decreased when the dose was divided. The only statistically significant correlations between the percentage of survival of the mice after 6 days and each of the PK/PD parameters were for peak concentration (Cmax)/MIC andS. aureusand for the free fraction ofCmax(Cmax-free)/MIC andS. pneumoniae. ForS. pneumoniae, the ED50for different dosing regimens increased with the number of doses given; e.g., the single-dose ED50s for vancomycin and teicoplanin were 0.65 and 0.45 mg/kg, respectively, but the ED50s for dosing regimens with 2-h doses given for 48 h were 6.79 and 5.67 mg/kg, respectively. In experiments with 39 different vancomycin dosing regimens and 40 different teicoplanin dosing regimens againstS. pneumoniae, the different PK/PD parameters were analyzed using logistic regression. TheCmax-free/MIC was one of two parameters that best explained the effect for both drugs; for vancomycin, the other important parameter was the AUC/MIC, and for teicoplanin, the other parameter was the time the free fraction of the drug is above the MIC. The effect analyzed as a function ofCmax-free/MIC disclosed thresholds with shifts from almost no effect to full effect at ratios of five to six for vancomycin and two to three for teicoplanin.
Spectinomycin Resistance in
Neisseria
spp. Due to Mutations in 16S rRNA
Authors: Galimand Marc , Gerbaud Guy , Courvalin Patrice 
Published: July 2002

ABSTRACT

Spectinomycin resistance in clinical isolates of
Neisseria meningitidis
and
Neisseria gonorrhoeae
was found to be due to mutations G1064C and C1192U (
Escherichia coli
numbering) in 16S rRNA genes, respectively.