bluuebunny/crossref_metadata_2025_split huggingface dataset:
Construction of a sequence motif characteristic of aminergic G protein–coupled receptors
Authors: Huang Enoch S. 
Published: June 2003

AbstractAn approach to discover sequence patterns characteristic of ligand classes is described and applied to aminergic G protein–coupled receptors (GPCRs). Putative ligand‐binding residue positions were inferred from considering three lines of evidence: conservation in the subfamily absent or underrepresented in the superfamily, any available mutation data, and the physicochemical properties of the ligand. For aminergic GPCRs, the motif is composed of a conserved aspartic acid in the third transmembrane (TM) domain (rhodopsin position 117) and a conserved tryptophan in the seventh TM domain (rhodopsin position 293); the roles of each are readily justified by molecular modeling of ligand‐receptor interactions. This minimally defined motif is an appropriate computational tool for identifying additional, potentially novel aminergic GPCRs from a set of experimentally uncharacterized “orphan” GPCRs, complementing existing sequence matching, clustering, and machine‐learning techniques. Motif sensitivity stems from the stepwise addition of residues characteristic of an entire class of ligand (and not tailored for any particular biogenic amine). This sensitivity is balanced by careful consideration of residues (evidence drawn from mutation data, correlation of ligand properties to residue properties, and location with respect to the extracellular face), thereby maintaining specificity for the aminergic class. A number of orphan GPCRs assigned to the aminergic class by this motif were later discovered to be a novel subfamily of trace amine GPCRs, as well as the successful classification of the histamine H4 receptor.
Effects of denaturants and substitutions of hydrophobic residues on backbone dynamics of denatured staphylococcal nuclease
Authors: Ohnishi Satoshi , Shortle David 
Published: June 2003

AbstractAnalysis of residual dipolar couplings (RDCs) in the Δ131Δ fragment of staphylococcal nuclease has demonstrated that its ensemble‐averaged structure is resistant to perturbations such as high concentrations of urea, low pH, and substitution of hydrophobic residues, suggesting that its residual structure is encoded by local side‐chain/backbone interactions. In the present study, the effects of these same perturbations on the backbone dynamics of Δ131Δ were examined through 1H‐15N relaxation methods. Unlike the global structure reported by RDCs, the transverse relaxation rates R2 were quite sensitive to denaturing conditions. At pH 5.2, Δ131Δ exhibits an uneven R2 profile with several characteristic peaks involving hydrophobic chain segments. Protonation of carboxyl side chains by lowering the pH reduces the values of R2 along the entire chain, yet these characteristic peaks remain. In contrast, high concentrations of urea or the substitution of 10 hydrophobic residues eliminates these peaks and reduces the R2 values by a greater amount. The combination of low pH and high urea leads to further decreases in R2. These denaturant‐induced increases in backbone mobility are also reflected in decreases in 15N NOEs and in relaxation interference parameters, with the former reporting an increase in fast motions and the latter a decrease in slow motions. Comparison between the changes in chain dynamics and the corresponding changes in Stokes radius and the patterns of RDCs suggests that regional variations in backbone dynamics in denatured nuclease arise primarily from local contacts between hydrophobic side chains and local interactions involving charged carboxyl groups.
Recombinant production and solution structure of PcTx1, the specific peptide inhibitor of ASIC1a proton‐gated cation channels
Authors: Escoubas Pierre , Bernard Cédric , Lambeau Gérard , Lazdunski Michel , Darbon Hervé 
Published: June 2003

AbstractAcid‐sensing ion channels (ASICs) are thought to be important ion channels, particularly for the perception of pain. Some of them may also contribute to synaptic plasticity, learning, and memory. Psalmotoxin 1 (PcTx1), the first potent and specific blocker of the ASIC1a proton‐sensing channel, has been successfully expressed in the Drosophila melanogaster S2 cell recombinant expression system used here for the first time to produce a spider toxin. The recombinant toxin was identical in all respects to the native peptide, and its three‐dimensional structure in solution was determined by means of 1H 2D NMR spectroscopy. Surface characteristics of PcTx1 provide insights on key structural elements involved in the binding of PcTx1 to ASIC1a channels. They appear to be localized in the β‐sheet and the β‐turn linking the strands, as indicated by electrostatic anisotropy calculations, surface charge distribution, and the presence of residues known to be implicated in channel recognition by other inhibitor cystine knot (ICK) toxins.
Solution structure of Vibrio cholerae protein VC0424: A variation of the ferredoxin‐like fold
Authors: Ramelot Theresa A. , Ni Shuisong , Goldsmith‐Fischman Sharon , Cort John R. , Honig Barry , Kennedy Michael A. 
Published: June 2003

AbstractThe structure of Vibrio cholerae protein VC0424 was determined by NMR spectroscopy. VC0424 belongs to a conserved family of bacterial proteins of unknown function (COG 3076). The structure has an α‐β sandwich architecture consisting of two layers: a four‐stranded antiparallel β‐sheet and three side‐by‐side α‐helices. The secondary structure elements have the order αβαββαβ along the sequence. This fold is the same as the ferredoxin‐like fold, except with an additional long N‐terminal helix, making it a variation on this common motif. A cluster of conserved surface residues on the β‐sheet side of the protein forms a pocket that may be important for the biological function of this conserved family of proteins.
Structure and properties of recombinant human pyridoxine 5′‐phosphate oxidase
Authors: Musayev Faik N. , Di Salvo Martino L. , Ko Tzu‐Ping , Schirch Verne , Safo Martin K. 
Published: June 2003

AbstractPyridoxine 5′‐phosphate oxidase catalyzes the terminal step in the synthesis of pyridoxal 5′‐phosphate. The cDNA for the human enzyme has been cloned and expressed in Escherichia coli. The purified human enzyme is a homodimer that exhibits a low catalytic rate constant of ∼0.2 sec−1 and Km values in the low micromolar range for both pyridoxine 5′phosphate and pyridoxamine 5′‐phosphate. Pyridoxal 5′‐phosphate is an effective product inhibitor. The three‐dimensional fold of the human enzyme is very similar to those of the E. coli and yeast enzymes. The human and E. coli enzymes share 39% sequence identity, but the binding sites for the tightly bound FMN and substrate are highly conserved. As observed with the E. coli enzyme, the human enzyme binds one molecule of pyridoxal 5′‐phosphate tightly on each subunit.
The role of side chain conformational flexibility in surface recognition by Tenebrio molitor antifreeze protein
Authors: Daley Margaret E. , Sykes Brian D. 
Published: June 2003

AbstractTwo‐dimensional nuclear magnetic resonance spectroscopy was used to investigate the flexibility of the threonine side chains in the β‐helical Tenebrio molitor antifreeze protein (TmAFP) at low temperatures. From measurement of the 3Jαβ 1H‐1H scalar coupling constants, the χ1 angles and preferred rotamer populations can be calculated. It was determined that the threonines on the ice‐binding face of the protein adopt a preferred rotameric conformation at near freezing temperatures, whereas the threonines not on the ice‐binding face sample many rotameric states. This suggests that TmAFP maintains a preformed ice‐binding conformation in solution, wherein the rigid array of threonines that form the AFP‐ice interface matches the ice crystal lattice. A key factor in binding to the ice surface and inhibition of ice crystal growth appears to be the close surface‐to‐surface complementarity between the AFP and crystalline ice, and the lack of an entropic penalty associated with freezing out motions in a flexible ligand.
Structural analysis of UBL5, a novel ubiquitin‐like modifier
Authors: McNally Teresa , Huang Qiulong , Janis Richard S. , Liu Zhihong , Olejniczak Edward T. , Reilly Regina M. 
Published: June 2003

AbstractUBL5 is a widely expressed human protein that is strongly conserved across phylogeny. Orthologs of UBL5 occur in every eukaryotic genome characterized to date. The yeast ortholog of UBL5, HUB1, was reported to be a ubiquitin‐like protein modifier important for modulation of protein function. However, unlike ubiquitin and all other ubiquitin‐like modifiers, UBL5 and its yeast ortholog HUB1 both contain a C‐terminal di‐tyrosine motif followed by a single variable residue instead of the characteristic di‐glycine found in all other ubiquitin‐like modifiers. Here we describe the three‐dimensional structure of UBL5 determined by NMR. The overall structure of the protein was found to be very similar to ubiquitin despite the low ∼25% residue similarity. The signature C‐terminal di‐tyrosine residues in UBL5 are involved in the final β sheet of the protein. This is very different to the di‐glycine motif found in ubiquitin, which extends beyond the final β sheet. In addition, we have confirmed an earlier report of an interaction between UBL5 and the cyclin‐like kinase, CLK4, which we have determined is specific and does not extend to other cyclin‐like kinase family members.
Biosynthetic Origin of Hygromycin A
Authors: Habib El-Sayed E. , Scarsdale J. Neel , Reynolds Kevin A. 
Published: June 2003

ABSTRACTHygromycin A, an antibiotic produced byStreptomyces hygroscopicus, is an inhibitor of bacterial ribosomal peptidyl transferase. The antibiotic binds to the ribosome in a distinct but overlapping manner with other antibiotics and offers a different template for generation of new agents effective against multidrug-resistant pathogens. Reported herein are the results from a series of stable-isotope-incorporation studies demonstrating the biosynthetic origins of the three distinct structural moieties which comprise hygromycin A. Incorporation of [1-13C]mannose and intact incorporation ofd-[1,2-13C2]glucose into the 6-deoxy-5-keto-d-arabino-hexofuranose moiety are consistent with a pathway in which mannose is converted to an activatedl-fucose, via a 4-keto-6-deoxy-d-mannose intermediate, with a subsequent unusual mutation of the pyranose to the corresponding furanose. The aminocyclitol moiety was labeled byd-[1,2-13C2]glucose in a manner consistent with formation ofmyo-inositol and a subsequent unprecedented oxidation and transamination of the C-2 hydroxyl group to generateneo-inosamine-2. Incorporation of [carboxy-13C]-4-hydroxybenzoic acid and intact incorporation of [2,3-13C2]propionate are consistent with a polyketide synthase-type decarboxylation condensation to generate the 3,4-dihydroxy-α-methylcinnamic acid moiety of hygromycin A. No labeling of hygromycin A was observed when [3-13C]tyrosine, [3-13C]phenylalanine, or [carboxy-13C]benzoic acid was used, suggesting that the 4-hydroxybenzoic acid is derived directly from chorismic acid. Consistent with this hypothesis was the observation that hygromycin A titers could be reduced by addition ofN-(phosphonomethyl)-glycine (an inhibitor of chorismic acid biosynthesis) and restored by coaddition of 4-hydroxybenzoic acid. The convergent biosynthetic pathway established for hygromycin A offers significant versatility for applying the techniques of combinatorial and directed biosynthesis to production of new antibiotics which target the ribosomal peptidyl transferase activity.
Viridans Group Streptococci Are Donors in Horizontal Transfer of Topoisomerase IV Genes to
Streptococcus pneumoniae
Authors: Balsalobre Luz , Ferrándiz María José , Liñares Josefina , Tubau Fe , de la Campa Adela G. 
Published: June 2003

ABSTRACT

A total of 46 ciprofloxacin-resistant (Cip
r
)
Streptococcus pneumoniae
strains were isolated from 1991 to 2001 at the Hospital of Bellvitge. Five of these strains showed unexpectedly high rates of nucleotide variations in the quinolone resistance-determining regions (QRDRs) of their
parC
,
parE
, and
gyrA
genes. The nucleotide sequence of the full-length
parC
,
parE
, and
gyrA
genes of one of these isolates revealed a mosaic structure compatible with an interspecific recombination origin. Southern blot analysis and nucleotide sequence determinations showed the presence of an
ant
-like gene in the intergenic
parE
-
parC
regions of the
S. pneumoniae
Cip
r
isolates with high rates of variations in their
parE
and
parC
QRDRs. The
ant
-like gene was absent from typical
S. pneumoniae
strains, whereas it was present in the intergenic
parE
-
parC
regions of the viridans group streptococci (
Streptococcus mitis
and
Streptococcus oralis
). These results suggest that the viridans group streptococci are acting as donors in the horizontal transfer of fluoroquinolone resistance genes to
S. pneumoniae
.
Burkholderia pseudomallei
Class A β-Lactamase Mutations That Confer Selective Resistance against Ceftazidime or Clavulanic Acid Inhibition
Authors: Tribuddharat Chanwit , Moore Richard A. , Baker Patricia , Woods Donald E. 
Published: June 2003

ABSTRACT

Burkholderia pseudomallei
, the causative agent of melioidosis, is inherently resistant to a variety of antibiotics including aminoglycosides, macrolides, polymyxins, and β-lactam antibiotics. Despite resistance to many β-lactams, ceftazidime and β-lactamase inhibitor-β-lactam combinations are commonly used for treatment of melioidosis. Here, we examine the enzyme kinetics of β-lactamase isolated from mutants resistant to ceftazidime and clavulanic acid inhibition and describe specific mutations within conserved motifs of the β-lactamase enzyme which account for these resistance patterns. Sequence analysis of regions flanking the
B. pseudomallei penA
gene revealed a putative regulator gene located downstream of
penA
. We have cloned and sequenced the
penA
gene from
B. mallei
and found it to be identical to
penA
from
B. pseudomallei
.
Extended-Spectrum Cephalosporin Compared to Cefazolin for Treatment of
Klebsiella pneumoniae
-Caused Liver Abscess
Authors: Cheng Hsiao-Pei , Siu L. K. , Chang Feng-Yee 
Published: June 2003

ABSTRACT

From January 1995 to May 2000, a total of 107 adults with liver abscess due to
Klebsiella pneumoniae
admitted at a large medical center in northern Taiwan were reviewed. Patients were considered to have received cefazolin or an extended-spectrum cephalosporin if they received at least 3 days of that antibiotic within the first 5 days of hospitalization. Fifty-nine (55.1%) patients received cefazolin, and 48 (44.9%) patients received an extended-spectrum cephalosporin. The demographic data, clinical features, severities of illness, and rates of early drainage for the two groups were comparable. However, the rates of developing complications for the two groups were significantly different (37.3 versus 6.3%, respectively;
P
< 0.001). Furthermore, six independent factors preventing severe complications following liver abscess due to
K. pneumoniae
were identified: normal platelet count, alkaline phosphatase less than 300 U/liter, no gas formation in the abscess, APACHE III score less than 40, use of an extended-spectrum cephalosporin, and early drainage. In conclusion, cefazolin therapy may be suboptimal for patients with liver abscess due to
K. pneumoniae
despite active in vitro susceptibility. Use of an extended-spectrum cephalosporin and early drainage for patients with liver abscess due to
K. pneumoniae
are suggested.
Correlation between Resistance ofPseudomonas aeruginosato Quaternary Ammonium Compounds and Expression of Outer Membrane Protein OprR
Authors: Tabata Atsushi , Nagamune Hideaki , Maeda Takuya , Murakami Keiji , Miyake Yoichiro , Kourai Hiroki 
Published: June 2003

ABSTRACTThe adaptation mechanism ofPseudomonas aeruginosaATCC 10145 to quaternary ammonium compounds (QACs) was investigated. AP. aeruginosastrain with adapted resistance to QACs was developed by a standard broth dilution method. It was revealed thatP. aeruginosaexhibited remarkable resistance toN-dodecylpyridinium iodide (P-12), whose structure is similar to that of a common disinfectant, cetylpyridinium chloride. Adapted resistance to benzalkonium chloride (BAC), which is commonly used as a disinfectant, was also observed inP. aeruginosa. Moreover, the P-12-resistant strain exhibited cross-resistance to BAC. Analysis of the outer membrane protein of the P-12-resistant strain by two-dimensional polyacrylamide gel electrophoresis showed a significant increase in the level of expression of a protein (named OprR) whose molecular mass was approximately 26 kDa. The actual function of OprR is not yet clear; however, OprR was expected to be an outer membrane-associated protein with homology to lipoproteins of other bacterial species, according to a search of the National Center for Biotechnology Information website with the BLAST program by use of the N-terminal sequence of OprR. A correlation between the level of expression of OprR and the level of resistance ofP. aeruginosato QACs was observed by using a PA2800 gene knockout mutant derived from the P-12-resistant strain. The knockout mutant recovered susceptibility not only to P-12 but also to BAC. These results suggested that OprR significantly participated in the adaptation ofP. aeruginosato QACs, such as P-12 and BAC.
Influences of Methodological Variables on Susceptibility Testing of Caspofungin against
Candida
Species and
Aspergillus fumigatus
Authors: Bartizal C. , Odds F. C. 
Published: June 2003

ABSTRACT

The influences of test variables on the outcome of susceptibility testing with caspofungin were tested with isolates of
Candida
spp. and
Aspergillus fumigatus
. Among six growth conditions tested with a range of inoculum sizes, the highest control growth yields were obtained in Sabouraud broth for all fungi, followed by RPMI 1640 (pH 7) for
Candida
spp. and antibiotic medium 3 (AM3) for
A. fumigatus
. RPMI 1640 gave unacceptably low growth yields with
A. fumigatus
. The caspofungin MICs under these various conditions ranged over more than 4 twofold dilutions for 7 of 16 fungi tested when a 50% inhibition (50% inhibitory concentration [IC
50
]) endpoint was used and for 12 of 16 fungi tested when an 80% inhibition (IC
80
) endpoint was used. A multifactorial design to study the influences of six test variables on control growth and the MIC showed that, for 14 isolates of
Candida
spp., the glucose concentration and the medium composition were the most common factors significantly influencing both control growth yields and the MIC. For eight
A. fumigatus
isolates, incubation time (24 versus 48 h) and temperature (30 versus 35°C) significantly affected control optical density (OD) values, while growth medium (AM3 versus Sabouraud broth) was the most common process variable affecting the MICs. Tests with AM3 from three suppliers showed significant variations in control OD values related to supplier, but IC
50
s fell within a 2- or 3-dilution range for 19 (86%) of the 22 isolates tested. We recommend that, at present, AM3 is superior to RPMI 1640 for testing of the susceptibilities of both yeasts and filamentous fungi to caspofungin and that a minimum incubation time of 48 h is necessary to test
A. fumigatus
adequately.
Chemosensitization of
Plasmodium falciparum
by Probenecid In Vitro
Authors: Nzila Alexis , Mberu Eddy , Bray Pat , Kokwaro Gilbert , Winstanley Peter , Marsh Kevin , Ward Steve 
Published: June 2003

ABSTRACT

Resistance to drugs can result from changes in drug transport, and this resistance can sometimes be overcome by a second drug that modifies the transport mechanisms of the cell. This strategy has been exploited to partly reverse resistance to chloroquine in
Plasmodium falciparum
. Studies with human tumor cells have shown that probenecid can reverse resistance to the antifolate methotrexate, but the potential for reversal of antifolate resistance has not been studied in
P. falciparum.
In the present study we tested the ability of probenecid to reverse antifolate resistance in
P. falciparum
in vitro. Probenecid, at concentrations that had no effect on parasite viability alone (50 μM), was shown to increase the sensitivity of a highly resistant parasite isolate to the antifolates pyrimethamine, sulfadoxine, chlorcycloguanil, and dapsone by seven-, five-, three-, and threefold, respectively. The equivalent effects against an antifolate-sensitive isolate were activity enhancements of approximately 3-, 6-, 1.2-, and 19-fold, respectively. Probenecid decreased the level of uptake of radiolabeled folic acid, suggesting a transport-based mechanism linked to folate salvage. When probenecid was tested with chloroquine, it chemosensitized the resistant isolate to chloroquine (i.e., enhanced the activity of chloroquine). This enhancement of activity was associated with increased levels of chloroquine accumulation. In conclusion, we have shown that probenecid can chemosensitize malaria parasites to antifolate compounds via a mechanism linked to reduced folate uptake. Notably, this effect is observed in both folate-sensitive and -resistant parasites. In contrast to the activities of antifolate compounds, the effect of probenecid on chloroquine sensitivity was selective for chloroquine-resistant parasites (patent P407595GB [W. P. Thompson & Co., Liverpool, United Kingdom] has been filed to protect this intellectual property).
Synergistic Antimicrobial Activity of Metabolites Produced by a Nonobligate Bacterial Predator
Authors: Cain Cody C. , Lee Dongho , Waldo Robert H. , Henry Alexis T. , Casida Earl J. , Wani Mansukh C. , Wall Monroe E. , Oberlies Nicholas H. , Falkinham Joseph O. 
Published: June 2003

ABSTRACT

A naturally occurring, gram-negative, nonobligate predator bacterial strain 679-2, exhibits broad-spectrum antimicrobial activity that is due, in part, to the production of three extracellular compounds. Antimicrobial-activity-directed fractionation of a culture of strain 679-2 against a panel of microorganisms has led to the isolation of three compounds: pyrrolnitrin, maculosin, and a new compound, which we have named banegasine. Although pyrrolnitrin is well known in the literature, it has not been found in cells with the herbicide maculosin. Further, this is the first report of production of maculosin by a prokaryote. Both maculosin and banegasine, which displayed no antimicrobial activities alone, were found to potentiate the antimicrobial activity of pyrrolnitrin. Based on 16S rRNA sequence, cellular fatty acid composition, and biochemical and cultural characteristics, strain 679-2 appears to represent a new genus and species of eubacteria,
Aristabacter necator
. The potent, broad-spectrum antimicrobial activity of predator strain 679-2 may be due to synergism between metabolites.