bluuebunny/crossref_metadata_2025_split huggingface dataset:
Proteins of circularly permuted sequence present within the same organism: The major serine proteinase inhibitor from Capsicum annuum seeds
Authors: Antcheva Nikolinka , Pintar Alessandro , Patthy András , Simoncsits András , Barta Endre , Tchorbanov Bojidar , Pongor Sándor 
Published: July 2002

AbstractThe major serine proteinase inhibitor from bell pepper (Capsicum annuum, paprika) seeds was isolated, characterized, and sequenced, and its disulfide bond topology was determined. PSI‐1.2 is a 52‐amino‐acid‐long, cysteine‐rich polypeptide that inhibits both trypsin (Ki = 4.6 × 10−9 M) and chymotrypsin (Ki = 1.1 × 10−8 M) and is a circularly permuted member of the potato type II inhibitor family. Mature proteins of this family are produced from precursor proteins containing two to eight repeat units that are proteolytically cleaved within, rather than between, the repeats. In contrast, PSI‐1.2 corresponds to a complete repeat that was predicted as the putative ancestral protein of the potato type II family. To our knowledge, this is the first case in which two proteins related to each other by circular permutation are shown to exist in the same organism and are expressed within the same organ. PSI‐1.2 is not derived from any of the known precursors, and it contains a unique amphiphilic segment in one of its loops. A systematic comparison of the related precursor repeat‐sequences reveals common evolutionary patterns that are in agreement with the ancestral gene‐duplication hypothesis.
Role of conserved residues in structure and stability: Tryptophans of human serum retinol‐binding protein, a model for the lipocalin superfamily
Authors: Greene Lesley H. , Chrysina Evangelia D. , Irons Laurence I. , Papageorgiou Anastassios C. , Acharya K. Ravi , Brew Keith 
Published: July 2002

AbstractSerum retinol binding protein (RBP) is a member of the lipocalin family, proteins with up‐and‐down β‐barrel folds, low levels of sequence identity, and diverse functions. Although tryptophan 24 of RBP is highly conserved among lipocalins, it does not play a direct role in activity. To determine if Trp24 and other conserved residues have roles in stability and/or folding, we investigated the effects of conservative substitutions for the four tryptophans and some adjacent residues on the structure, stability, and spectroscopic properties of apo‐RBP. Crystal structures of recombinant human apo‐RBP and of a mutant with substitutions for tryptophans 67 and 91 at 1.7 Å and 2.0 Å resolution, respectively, as well as stability measurements, indicate that these relatively exposed tryptophans have little influence on structure or stability. Although Trp105 is largely buried in the wall of the β‐barrel, it can be replaced with minor effects on stability to thermal and chemical unfolding. In contrast, substitutions of three different amino acids for Trp24 or replacement of Arg139, a conserved residue that interacts with Trp24, lead to similar large losses in stability and lower yields of native protein generated by in vitro folding. The results and the coordinated nature of natural substitutions at these sites support the idea that conserved residues in functionally divergent homologs have roles in stabilizing the native relative to misfolded structures. They also establish conditions for studies of the kinetics of folding and unfolding by ideying spectroscopic signals for monitoring the formation of different substructures.
Chemical‐modification rescue assessed by mass spectrometry demonstrates that γ‐thia‐lysine yields the same activity as lysine in aldolase
Authors: Hopkins Christopher E. , O'connor Peter B. , Allen Karen N. , Costello Catherine E. , Tolan Dean R. 
Published: July 2002

AbstractThe role of active site residues in fructose 1,6‐bisphosphate aldolase is investigated by chemical‐modification rescue. An active‐site mutation, K107C, is constructed in a background where the four solvent‐accessible cysteine residues are converted to alanine. The resulting mutant, tetK107C, when reacted with bromoethylamine (BrEA), shows a 40‐fold increase in activity (to 80% that of wild type). Determination of the sites and their degree of modification using electrospray ionization Fourier transform mass spectrometry (ESI‐FTMS) is developed, allowing correlation of activity after chemical modification rescue to the degree of modification. The stoichiometry of the reaction is 2.5 aminoethylations per subunit, as measured by ESI‐FTMS. Protein modification with a double‐labeled mix (1:1) of natural abundance isotope (d0‐BrEA) and 2‐bromoethyl‐1,1,2,2‐d4‐amine hydrobromide (d4‐BrEA), followed by dialysis and trypsin digestion, shows aminoethylated peptides as “twin peptides” separated by four mass units in ESI‐FTMS analysis. Using this detection procedure under nondenaturing (native) conditions, C107 is aminoethylated, whereas the four buried thiols remain unlabeled. Aminoethylation of other residues is observed, and correlates with those peptides containing histidine, methionine, and/or the amino terminus. Quantification of the aminoethylation reaction is achieved by labeling with nondeuterated d0‐BrEA under denaturing conditions following double labeling under native conditions. In addition to complete labeling all five thiols, the intensity of the d0‐BrEA peak for C107 containing peptides increases, and the change in the d0/d4 ratio between native and denaturing conditions shows 82 ± 4.5% aminoethylation at C107. This correlation of modification with the recovered activity, indicates that γ‐thia‐lysine replaces lysine in the catalytic mechanism. Kinetic constants measured for the rescued K107C mutant enzyme with the substrates fructose 1‐phosphate and fructose 1,6‐bisphosphate are consistent with the role of the positively charged lysine binding to the C6‐phosphate. ESI‐FTMS, combined with this double‐labeling procedure, allows precise identification of sites and measurement of degree of protein modification.
On Transition to Turbulence and Synthesis of Alternative Approaches
Authors: Lin C. C. 
Published: July 2002

In the study of complex systems, controversial debates often arise among advocates of different schools of thought. In this article, we examine how such controversies should be addressed, with the problem of transition to turbulence as a primary example.It is shown that, in many cases, these controversies may be resolved by first noting that the alternative theories proposed may not be mutually exclusive. Indeed, they may even be mutually complementary, if they were originally developed to address similar issues in different physical contexts. In any case, for the validity of the alternative theories proposed, each should be separately and fully supported from both the theoretical and empirical points of view. Each applies to a specific physical context, and each stands on its own merits and limitations. Synthesis into a broader theory may then be achieved, if commonality is identified among the different alternative theories proposed.To demonstrate this conciliatory approach, we begin with an examination of the move toward resolution of the well‐known controversy over the problem of transition to turbulence from the steady laminar flow in the boundary layer over a flat plate. Several other long‐standing controversies have been successfully addressed on the basis of this approach. In addition to the problem of transition to turbulence, we considered, in some detail, two additional examples: (1) the global structures of spiral galaxies; and (2) the theory of jet noise. In all three cases, it is shown that the model approach is meritorious despite the limitations.Synthesis, with a conciliatory approach to apparent conflicts, will be recommended in general as a new part of an extended paradigm in applied mathematics. It is an approach appropriate to situations where an ideal theory, with universal applicability, is elusive. Parallel development of several alternative theories is natural, and a final synthesis is needed. In contrast, it should be noted that the same perspective is generally not expected useful if the controversies concern the unique solution of well‐defined mathematical issues. The potential success of the application of this conciliatory perception and approach to other areas of science are discussed (see Section 5).
In Vitro Increase in Chloroquine Accumulation Induced by Dihydroethano- and Ethenoanthracene Derivatives in
Plasmodium falciparum
-Parasitized Erythrocytes
Authors: Pradines Bruno , Alibert Sandrine , Houdoin Carole , Santelli-Rouvier Christiane , Mosnier Joel , Fusai Thierry , Rogier Christophe , Barbe Jacques , Parzy Daniel 
Published: July 2002

ABSTRACT
The effects of a series of dihydroethano- and ethenoanthracene derivatives on chloroquine (CQ) accumulation in CQ-susceptible strain 3D7 and CQ-resistant clone W2 were assessed. The levels of CQ accumulation increased little or none in CQ-susceptible strain 3D7 and generally increased markedly in CQ-resistant strain W2. At 10 μM, 28 compounds yielded cellular accumulation ratios (CARs) greater than that observed with CQ alone in W2. At 10 μM, in strain W2, 21 of 31 compounds had CQ CARs two or more times higher than that of CQ alone, 15 of 31 compounds had CQ CARs three or more times higher than that of CQ alone, 13 of 31 compounds had CQ CARs four or more times higher than that of CQ alone, and 9 of 31 compounds had CQ CARs five or more times higher than that of CQ alone. At 1 μM, 17 of 31 compounds had CQ CARs two or more times higher than that of CQ alone, 12 of 31 compounds had CQ CARs three or more times higher than that of CQ alone, 6 of 31 compounds had CQ CARs four or more times higher than that of CQ alone, and 3 of 31 compounds had CQ CARs five or more times higher than that of CQ alone. At 1 μM, 17 of 31 compounds were more potent inducers of CQ accumulation than verapamil and 12 of 31 compounds were more potent inducers of CQ accumulation than promethazine. The nature of the basic group seems to be associated with increases in the levels of CQ accumulation. At 1 and 10 μM, 10 of 14 and 13 of 14 compounds with amino group (amines and diamines), respectively, had CARs ≥3, while at 1 and 10 μM, only 1 of the 13 derivatives with amido groups had CARs ≥3. Among 12 of the 31 compounds which were more active inducers of CQ accumulation than promethazine at 1 μM, 10 had amino groups and 1 had an amido group.
Comparative Intracellular (THP-1 Macrophage) and Extracellular Activities of β-Lactams, Azithromycin, Gentamicin, and Fluoroquinolones against
Listeria monocytogenes
at Clinically Relevant Concentrations
Authors: Carryn Stéphane , Van Bambeke Françoise , Mingeot-Leclercq Marie-Paule , Tulkens Paul M. 
Published: July 2002

ABSTRACT

The activities of ampicillin, meropenem, azithromycin, gentamicin, ciprofloxacin, and moxifloxacin against intracellular hemolysin-positive
Listeria monocytogenes
were measured in human THP-1 macrophages and were compared with the extracellular activities observed in broth. All extracellular concentrations were adjusted to explore ranges that are clinically achievable in human serum upon conventional therapy. In broth, ampicillin, meropenem, and azithromycin were only bacteriostatic, whereas gentamicin, ciprofloxacin, and moxifloxacin were strongly bactericidal in a concentration-dependent manner. In cells, ampicillin, meropenem, azithromycin, and ciprofloxacin were slightly bactericidal (0.3- to 0.8-log CFU reductions), moxifloxacin was strongly bactericidal (2.1-log CFU reduction), and gentamicin was virtually inactive. The difference in the efficacies of moxifloxacin and ciprofloxacin in cells did not result from a difference in levels of accumulation in cells (6.96 ± 1.05 versus 7.75 ± 1.03) and was only partially explainable by the difference in the MICs (0.58 ± 0.04 versus 1.40 ± 0.17 mg/liter). Further analysis showed that intracellular moxifloxacin expressed only approximately 1/7 of the activity demonstrated against extracellular bacteria and ciprofloxacin expressed only 1/15 of the activity demonstrated against extracellular bacteria. Gentamicin did not increase the intracellular activities of the other antibiotics tested. The data suggest (i) that moxifloxacin could be of potential interest for eradication of the intracellular forms of
L. monocytogenes
, (ii) that the cellular accumulation of an antibiotic is not the only determinant of its intracellular activity (for fluoroquinolones, it is actually a self-defeating process as far as activity is concerned), and (iii) that pharmacodynamics (activity-to-concentration relationships) need to be considered for the establishment of efficacy against intracellular bacteria, just as they are for the establishment of efficacy against extracellular infections.
CmeABC Functions as a Multidrug Efflux System in
Campylobacter jejuni
Authors: Lin Jun , Michel Linda Overbye , Zhang Qijing 
Published: July 2002

ABSTRACT

Campylobacter jejuni
, a gram-negative organism causing gastroenteritis in humans, is increasingly resistant to antibiotics. However, little is known about the drug efflux mechanisms in this pathogen. Here we characterized an efflux pump encoded by a three-gene operon (designated
cmeABC
) that contributes to multidrug resistance in
C. jejuni
81-176. CmeABC shares significant sequence and structural homology with known tripartite multidrug efflux pumps in other gram-negative bacteria, and it consists of a periplasmic fusion protein (CmeA), an inner membrane efflux transporter belonging to the resistance-nodulation-cell division superfamily (CmeB), and an outer membrane protein (CmeC). Immunoblotting using CmeABC-specific antibodies demonstrated that
cmeABC
was expressed in wild-type 81-176; however, an isogenic mutant (9B6) with a transposon insertion in the
cmeB
gene showed impaired production of CmeB and CmeC. Compared to wild-type 81-176, 9B6 showed a 2- to 4,000-fold decrease in resistance to a range of antibiotics, heavy metals, bile salts, and other antimicrobial agents. Accumulation assays demonstrated that significantly more ethidium bromide and ciprofloxacin accumulated in mutant 9B6 than in wild-type 81-176. Addition of carbonyl cyanide
m
-chlorophenylhydrazone, an efflux pump inhibitor, increased the accumulation of ciprofloxacin in wild-type 81-176 to the level of mutant 9B6. PCR and immunoblotting analysis also showed that
cmeABC
was broadly distributed in various
C. jejuni
isolates and constitutively expressed in wild-type strains. Together, these findings formally establish that CmeABC functions as a tripartite multidrug efflux pump that contributes to the intrinsic resistance of
C. jejuni
to a broad range of structurally unrelated antimicrobial agents.
Resistance to Quinupristin-Dalfopristin Due to Mutation of L22 Ribosomal Protein inStaphylococcus aureus
Authors: Malbruny Brigitte , Canu Annie , Bozdogan Bülent , Fantin Bruno , Zarrouk Virginie , Dutka-Malen Sylvie , Feger Celine , Leclercq Roland 
Published: July 2002

ABSTRACTThe mechanism of resistance to the streptogramin antibiotics quinupristin and dalfopristin was studied in aStaphylococcus aureusclinical isolate selected under quinupristin-dalfopristin therapy, in four derivatives ofS. aureusRN4220 selected in vitro, and in a mutant selected in a model of rabbit aortic endocarditis. For all strains the MICs of erythromycin, quinupristin, and quinupristin-dalfopristin were higher than those for the parental strains but the MICs of dalfopristin and lincomycin were similar. Portions of genes for domains II and V of 23S rRNA and the genes for ribosomal proteins L4 and L22 were amplified and sequenced. All mutants contained insertions or deletions in a protruding β hairpin that is part of the conserved C terminus of the L22 protein and that interacts with 23S rRNA. SusceptibleS. aureusRN4220 was transformed with plasmid DNA encoding the L22 alteration, resulting in transformants that were erythromycin and quinupristin resistant. Synergistic ribosomal binding of streptogramins A and B, studied by analyzing the fluorescence kinetics of pristinamycin IA-ribosome complexes, was abolished in the mutant strain, providing an explanation for quinupristin-dalfopristin resistance.
Acyclovir Cream for Treatment of Herpes Simplex Labialis: Results of Two Randomized, Double-Blind, Vehicle-Controlled, Multicenter Clinical Trials
Authors: Spruance Spotswood L. , Nett Robert , Marbury Thomas , Wolff Ray , Johnson James , Spaulding for The Acyclovir Cream Study Group Theodore 
Published: July 2002

ABSTRACT

Acyclovir cream has been available for the treatment of herpes labialis in numerous countries outside the United States for over a decade. Evidence for its efficacy comes from a few small clinical trials conducted in the 1980s. To examine more comprehensively the efficacy and safety of this formulation, we conducted two independent, identical, parallel, randomized, double-blind, vehicle-controlled, large-scale multicenter clinical trials. Healthy adults with a history of frequent herpes labialis were recruited from the general population, screened for eligibility, randomized equally to 5% acyclovir cream or vehicle control, given study medication, and told to self-initiate treatment five times daily for 4 days beginning within 1 h of the onset of a recurrent episode. The number of patients who treated a lesion was 686 in study 1 and 699 in study 2. In study 1, the mean duration of episodes was 4.3 days for patients treated with acyclovir cream and 4.8 days for those treated with the vehicle control (hazards ratio [HR] = 1.23; 95% confidence interval [CI], 1.06 to 1.44;
P
= 0.007). In study 2, the mean duration of episodes was 4.6 days for patients treated with acyclovir cream and 5.2 days for those treated with the vehicle control (HR = 1.24; 95% CI, 1.06 to 1.44;
P
= 0.006). Efficacy was apparent whether therapy was initiated “early” (prodrome or erythema lesion stage) or “late” (papule or vesicle stage). There was a statistically significant reduction in the duration of lesion pain in both studies. Acyclovir cream did not prevent the development of classical lesions (progression to vesicles, ulcers, and/or crusts). Adverse events were mild and infrequent.
Inhibition of Penicillinase by Epigallocatechin Gallate Resulting in Restoration of Antibacterial Activity of Penicillin against Penicillinase-Producing
Staphylococcus aureus
Authors: Zhao Wei-Hua , Hu Zhi-Qing , Hara Yukihiko , Shimamura Tadakatsu 
Published: July 2002

ABSTRACT

The combination of epigallocatechin gallate (EGCg, a main constituent of tea catechins) with penicillin showed synergism against 21 clinical isolates of penicillinase-producing
Staphylococcus aureus
. Besides binding directly to peptidoglycan, the inhibition of penicillinase activity by EGCg is responsible for the synergism. EGCg inhibited the penicillinase activity in a dose-dependent fashion, with a 50% inhibitory concentration of 10 μg/ml.
bla
CTX-M-2
Is Located in an Unusual Class 1 Integron (In35) Which Includes Orf513
Authors: Arduino Sonia M. , Roy Paul H. , Jacoby George A. , Orman Betina E. , Pineiro Silvia A. , Centron Daniela 
Published: July 2002

ABSTRACT

Examination of the
bla
CTX-M-2
gene in plasmid pMAR-12 by sequencing and PCR analysis revealed that the
bla
gene and the surrounding DNA, which is closely related (99% homology) to the
Kluyvera ascorbata
chromosomal DNA that contains the
bla
KLUA-1
gene, are located in a complex
sul1
-type integron, termed In35, that includes Orf513. It is possible that
bla
CTX-M-2
was acquired by plasmid pMAR-12 through an uncharacterized recombinational event in which Orf513 could be involved.
Two Diarylurea Electron Transport Inhibitors Reduce
Staphylococcus aureus
Hemolytic Activity and Protect Cultured Endothelial Cells from Lysis
Authors: Proctor R. A. , Dalal S. C. , Kahl B. , Brar D. , Peters G. , Nichols W. W. 
Published: July 2002

ABSTRACT

Reduction in electron transport is associated with decreased production in alpha-toxin despite the fact that
Staphylococcus aureus
is able to grow from 1 CFU to >10
7
CFU. Similarly, under anaerobic conditions,
S. aureus
does not produce alpha-toxin. Although the pathways that connect oxidative metabolism and toxin production are unknown, agents are available that exhibit greater inhibition of plant versus mammalian electron transport. Herbicides block electron transport in plants by inhibiting the formation of phosphoquinol (QH
2
) in plants. Commercial use in farming is possible because these compounds are much less active against the quinones found mammalian mitochondria. Because bacterial electron transport systems are closer to plant than mammalian systems, we hypothesized that inhibitors of respiration might be able to reduce
S. aureus
electron transport and block the production of alpha-toxin. We studied two compounds and found that the effective dose for the inhibition of bacterial respiration was 50 to >3,500 times lower than the concentration required to cause similar inhibition of rat mitochondrial respiration. Compounds I and II also reduced toxin production in
S. aureus
without causing overt toxicity to cultured endothelial cells. Finally, the compounds reduced the damage caused by
S. aureus
when cocultured with the endothelial cells. This raises the possibility that compounds that inhibit bacterial respiration might be prove valuable for the prevention of toxin production in
S. aureus
.
Steady-State Pharmacokinetics of Lamivudine in Human Immunodeficiency Virus-Infected Patients with End-Stage Renal Disease Receiving Chronic Dialysis
Authors: Bohjanen Paul R. , Johnson Melissa D. , Szczech Lynda A. , Wray Dannah W. , Petros William P. , Miller Cameron R. , Hicks Charles B. 
Published: July 2002

ABSTRACT

The steady-state pharmacokinetics of lamivudine were evaluated in 11 subjects with human immunodeficiency virus infection and end-stage renal disease, 9 of whom were receiving hemodialysis and 2 of whom were receiving chronic ambulatory peritoneal dialysis (CAPD). All subjects received 150 mg of lamivudine daily for at least 2 weeks prior to sampling for determination of the pharmacokinetics of lamivudine over a 24-h period on 2 consecutive days. On the first day, subjects received 150 mg of oral lamivudine and underwent dialysis (hemodialysis or CAPD). On the second day, subjects received another 150 mg of oral lamivudine but dialysis was not performed. For the subjects undergoing hemodialysis, the geometric mean predose serum lamivudine concentration was 1.14 μg/ml (95% confidence interval [CI], 0.83 to 1.58 μg/ml), the geometric mean maximum concentration in serum (
C
max
) was 3.77 μg/ml (95% CI, 3.01 to 4.71 μg/ml), and the geometric mean area under the serum concentration-time curve from time zero to 24 h (AUC
0-24
) was 49.8 μg · h/ml (95% CI 39.1 to 63.6 μg · h/ml). Hemodialysis removed approximately 28 mg of lamivudine but had no significant effect on
C
max
or AUC
0-24
. In the absence of hemodialysis, the geometric mean lamivudine terminal elimination half-life was 17.2 h (95% CI, 10.5 to 28.1 h), whereas the geometric mean intradialysis half-life of lamivudine was 5.3 h (95% CI, 3.4 to 8.2 h). The pharmacokinetics of lamivudine in subjects undergoing CAPD were similar to those in subjects undergoing hemodialysis. CAPD removed 24 mg of lamivudine over a 24-h period but had no effect on
C
max
or AUC
0-24
. Pharmacokinetic modeling suggests that a lamivudine dose of 25 mg daily in hemodialysis subjects would provide serum exposure similar to that provided by a dose of 150 mg twice daily in patients with normal renal function.