bluuebunny/crossref_metadata_2025_split huggingface dataset:
Involvement of a Cytochrome P450 Monooxygenase in Thaxtomin A Biosynthesis by
Streptomyces acidiscabies
Authors: Healy F. G. , Krasnoff S. B. , Wach M. , Gibson D. M. , Loria R. 
Published: July 2002

ABSTRACT

The biosynthesis of the thaxtomin cyclic dipeptide phytotoxins proceeds nonribosomally via the thiotemplate mechanism. Acyladenylation, thioesterification,
N
-methylation, and cyclization of two amino acid substrates are catalyzed by the
txtAB
-encoded thaxtomin synthetase. Nucleotide sequence analysis of the region 3′ of
txtAB
in
Streptomyces acidiscabies
84.104 identified an open reading frame (ORF) encoding a homolog of the P450 monooxygenase gene family. It was proposed that thaxtomin A phenylalanyl hydroxylation was catalyzed by the monooxygenase homolog. The ORF was mutated in
S. acidiscabies
84.104 by using an integrative gene disruption construct, and culture filtrate extracts of the mutant were assayed for the presence of dehydroxy derivatives of thaxtomin A. Reversed-phase high-performance liquid chromatography (HPLC) and HPLC-mass spectrometry indicated that the major component in culture filtrate extracts of the mutant was less polar and smaller than thaxtomin A. Comparisons of electrospray mass spectra as well as
1
H- and
13
C-nuclear magnetic resonance spectra of the purified compound with those previously reported for thaxtomins confirmed the structure of the compound as 12,15-
N
-dimethylcyclo-(
l
-4-nitrotryptophyl-
l
-phenylalanyl), the didehydroxy analog of thaxtomin A. The ORF, designated
txtC
, was cloned and the recombinant six-His-tagged fusion protein produced in
Escherichia coli
and purified from cell extracts. TxtC produced in
E. coli
exhibited spectral properties similar to those of cytochrome P450-type hemoproteins that have undergone conversion to the catalytically inactive P420 form. Based on these properties and the high similarity of TxtC to other well-characterized P450 enzymes, we conclude that
txtC
encodes a cytochrome P450-type monooxygenase required for postcyclization hydroxylation of the cyclic dipeptide.
Use of an Enzyme Immunoassay Does Not Eliminate the Need To Analyze Multiple Stool Specimens for Sensitive Detection of
Giardia lamblia
Authors: Hanson Kevan L. , Cartwright Charles P. 
Published: July 2002

ABSTRACT

The relative sensitivities of a commercially available enzyme immunoassay (EIA) (ProSpecT
Giardia
; Alexon-Trend Inc., Ramsey, Minn.) and conventional ovum-and-parasite (O&P) examination for the detection of
Giardia lamblia
in preserved stool specimens were determined. Paired stool samples collected independently within a 7-day period from 103 patients were analyzed by both methods. A total of 54 specimens from 30 patients (18 asymptomatically infected with
G. lamblia
and 12 with symptoms consistent with intestinal giardiasis) were determined to be positive for
G. lamblia
, of which 48 (88.9%) were positive by microscopy and 52 (96.3%) were positive by EIA. Both specimens submitted were positive for
G. lamblia
by O&P examination for 66.7% (20 of 30) of the positive patients; for 26.7% (8 of 30) a single specimen was positive by O&P examination, and for 6.7% (2 of 30) of those determined to be infected with
G. lamblia
, both samples were negative by microscopy. The sensitivity of conventional O&P examination was somewhat higher in symptomatically infected individuals, with 75% (9 of 12) of patients in this category having
G. lamblia
detected in both samples, compared with 61% (11 of 18) of asymptomatic patients. A total of 24 positive patients (80%) had
G. lamblia
antigen detected by EIA in both submitted samples, 4 positive patients (13.3%) had one specimen positive by EIA, and the EIA was negative in both specimens from 2 infected individuals (6.5%), the sensitivity of EIA was substantially equivalent in asymptomatic and symptomatic individuals (77 versus 83% of patients with positive results on both specimens). Although the sensitivity of EIA for the detection of
G. lamblia
on a single stool specimen was somewhat higher than that of conventional O&P examination in symptomatic patients (83 versus 75%), in asymptomatic patients (77 versus 61%), and overall (80 versus 67%), examination of two specimens by either EIA or microscopy was necessary to achieve a diagnostic sensitivity of greater than 90%.
Rapid and Sensitive Oligonucleotide Ligation Assay for Detection of Mutations in Human Immunodeficiency Virus Type 1 Associated with High-Level Resistance to Protease Inhibitors
Authors: Beck Ingrid A. , Mahalanabis Madhumita , Pepper Gregory , Wright Amy , Hamilton Shannon , Langston Erika , Frenkel Lisa M. 
Published: July 2002

ABSTRACTA sensitive, specific, and high-throughput oligonucleotide ligation assay (OLA) for the detection of genotypic human immunodeficiency virus type 1 (HIV-1) resistance to Food and Drug Administration-approved protease inhibitors was developed and evaluated. This ligation-based assay uses differentially modified oligonucleotides specific for wild-type or mutant sequences, allowing sensitive and simple detection of both genotypes in a single well of a microtiter plate. Oligonucleotides were designed to detect primary mutations associated with high-level resistance to amprenavir, nelfinavir, indinavir, ritonavir, saquinavir, and lopinavir, including amino acid substitutions D30N, I50V, V82A/S/T, I84V, N88D, and L90M. Plasma HIV-1 RNA from 54 infected patients was amplified by reverse transcription-PCR and sequenced by using dideoxynucleotide chain terminators for evaluation of mutations associated with drug resistance. These same amplicons were genotyped by the OLA at positions 30, 50, 82, 88, 84, and 90 for a total of 312 codons. The sensitivity of detection of drug-resistant genotypes was 96.7% (87 of 90 mutant codons) in the OLA compared to 92.2% (83 of 90) in consensus sequencing, presumably due to the increased sensitivity of the OLA. The OLA detected genetic subpopulations more often than sequencing, detecting 30 mixtures of mutant and wild-type sequences and two mixtures of drug-resistant sequences compared to 15 detected by DNA sequencing. Reproducible and semiquantitative detection of the mutant and the wild-type genomes by the OLA was observed by analysis of wild-type and mutant plasmid mixtures containing as little as 5% of either genotype in a background of the opposite genome. This rapid, simple, economical, and highly sensitive assay provides a practical alternative to dideoxy sequencing for genotypic evaluation of HIV-1 resistance to antiretrovirals.
Interaction of Epstein-Barr Virus Nuclear Antigen Leader Protein (EBNA-LP) with HS1-Associated Protein X-1: Implication of Cytoplasmic Function of EBNA-LP
Authors: Kawaguchi Yasushi , Nakajima Kaori , Igarashi Mie , Morita Tomoko , Tanaka Michiko , Suzuki Mikiko , Yokoyama Akihiko , Matsuda Go , Kato Kentaro , Kanamori Mikiko , Hirai Kanji 
Published: July 2002

ABSTRACTEpstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) consists of W1W2 repeats and a unique C-terminal Y1Y2 domain and has been suggested to play an important role in EBV-induced transformation. To identify the cellular factors interacting with EBNA-LP, we performed a yeast two-hybrid screen, using EBNA-LP cDNA containing four W1W2 repeats as bait and an EBV-transformed human peripheral blood lymphocyte cDNA library as the source of cellular genes. Our results were as follows. (i) All three cDNAs in positive yeast colonies were found to encode the same cellular protein, HS1-associated protein X-1 (HAX-1), which is localized mainly in the cytoplasm and has been suggested to be involved in the regulation of B-cell signal transduction and apoptosis. (ii) Mutational analysis of EBNA-LP revealed that the association with HAX-1 is mediated by the W1W2 repeat domain. (iii) A purified chimeric protein consisting of glutathioneS-transferase fused to EBNA-LP specifically formed complexes with HAX-1 transiently expressed in COS-7 cells. (iv) When EBNA-LP and HAX-1 were coexpressed in COS-7 cells, EBNA-LP was specifically coimmunoprecipitated with HAX-1. (v) Careful cell fractionation experiments of an EBV-infected lymphoblastoid cell line revealed that EBNA-LP is localized in the cytoplasm as well as in the nucleus. (vi) When EBNA-LP containing four W1W2 repeats was expressed in COS-7 cells, EBNA-LP was detected mainly in the nucleus by immunofluorescence assay. Interestingly, when EBNA-LP containing a single W1W2 repeat was expressed in COS-7 cells, EBNA-LP was localized predominantly in the cytoplasm and was colocalized with HAX-1. These results indicate that EBNA-LP is in fact present and may have a significant function in the cytoplasm, possibly by interacting with and affecting the function of HAX-1.
Recessive Resistance in
Pisum sativum
and Potyvirus Pathotype Resolved in a Gene-for-Cistron Correspondence between Host and Virus
Authors: Johansen I. Elisabeth , Lund Ole Søgaard , Hjulsager Charlotte K. , Laursen Jesper 
Published: July 2002

ABSTRACT

Pea seed-borne mosaic potyvirus (PSbMV) isolates are divided into pathotypes P-1, P-2, and P-4 according to their infection profile on a panel of
Pisum sativum
lines.
P. sativum
PI 269818 is resistant to P-1 and P-2 isolates and is susceptible to P-4 isolates. Resistance to P-1 is inherited as a single recessive gene, denoted
sbm-1,
and the pathogenicity determinant has previously been mapped to the virus-coded protein VPg. In the cultivar Bonneville, a second recessive gene,
sbm-2,
confers specific resistance to P-2. By exchanging cistrons between a P-2 and a P-4 isolate, the P3-6k1 cistron was identified as the PSbMV host-specific pathogenicity determinant on Bonneville. Exchange of P3-6k1 did not affect infection on PI 269818, and infection of Bonneville was not altered by substitution of the VPg cistron, indicating that P3-6k1 and VPg are independent determinants of pathotype-specific infectivity. On PI 269818 the pathogenicity determinant of both P-1 and P-2 mapped to the N terminus of VPg. This suggests that VPg from the P-1 and P-2 isolates are functionally similar on this host and that resistance to P-1 and P-2 in PI 269818 may operate by the same mechanism. Identification of VPg–
sbm-1
and P3-6k1–
sbm-2
as independent pairs of genetic interactors between PSbMV and
P. sativum
provides a simple explanation of the three known pathotypes of PSbMV. Furthermore, analysis of β-glucuronidase-tagged P-2 virus indicated that
sbm-2
resistance affected an early step in infection, implying that the P3-6k1 region plays a critical role in potyvirus replication or cell-to-cell movement.
Reovirus ςNS Protein Is Required for Nucleation of Viral Assembly Complexes and Formation of Viral Inclusions
Authors: Becker Michelle M. , Goral Mehmet I. , Hazelton Paul R. , Baer Geoffrey S. , Rodgers Steven E. , Brown Earl G. , Coombs Kevin M. , Dermody Terence S. 
Published: July 2002

ABSTRACT

Progeny virions of mammalian reoviruses are assembled in the cytoplasm of infected cells at discrete sites termed viral inclusions. Studies of temperature-sensitive (
ts
) mutant viruses indicate that nonstructural protein ςNS and core protein μ2 are required for synthesis of double-stranded (ds) RNA, a process that occurs at sites of viral assembly. We used confocal immunofluorescence microscopy and
ts
mutant reoviruses to define the roles of ςNS and μ2 in viral inclusion formation. In cells infected with wild-type (wt) reovirus, ςNS and μ2 colocalize to large, perinuclear structures that correspond to viral inclusions. In cells infected at a nonpermissive temperature with ςNS-mutant virus
tsE320
, ςNS is distributed diffusely in the cytoplasm and μ2 is contained in small, punctate foci that do not resemble viral inclusions. In cells infected at a nonpermissive temperature with μ2-mutant virus
tsH11.2
, μ2 is distributed diffusely in the cytoplasm and the nucleus. However, ςNS localizes to discrete structures in the cytoplasm that contain other viral proteins and are morphologically indistinguishable from viral inclusions seen in cells infected with wt reovirus. Examination of cells infected with wt reovirus over a time course demonstrates that ςNS precedes μ2 in localization to viral inclusions. These findings suggest that viral RNA-protein complexes containing ςNS nucleate sites of viral replication to which other viral proteins, including μ2, are recruited to commence dsRNA synthesis.
Efficient Activation of Viral Genomes by Levels of Herpes Simplex Virus ICP0 Insufficient To Affect Cellular Gene Expression or Cell Survival
Authors: Hobbs William E. , Brough Douglas E. , Kovesdi Imre , DeLuca Neal A. 
Published: July 2002

ABSTRACT

Herpes simplex virus (HSV) ICP0 can effectively activate gene expression from otherwise silent promoters contained on persisting viral genomes. However, the expression of high levels of ICP0, as from ICP4
−
HSV type 1 (HSV-1) vectors, results in marked toxicity. We have analyzed the results of ICP0 expressed from an E1
−
E4
−
adenovirus vector (AdS.11E4ICP0) in which ICP0 expression is controlled from the endogenous adenoviral E4 promoter. In this system, the expression level of ICP0 was reduced more than 1,000-fold relative to the level of expression from HSV-1 vectors. This low level of ICP0 did not affect cellular division or greatly perturb cellular metabolism as assessed by gene expression array analysis comparing the effects of HSV and adenovirus vector strains. However, this amount of ICP0 was sufficient to quantitatively destroy ND10 structures as measured by promyelocytic leukemia immunofluorescence. The levels of adenovirus-expressed ICP0 were sufficient to activate quiescent viral genomes in
trans
and promote persistent transgene expression in
cis
. Moreover, infection of complementing cells with AdS.11E4ICP0 promoted viral growth and resulted in a 20-fold increase in the plaquing efficiency of
d
109, a virus defective for all five immediate-early genes. Thus, the low level expression of ICP0 from the E1
−
E4
−
adenovirus vector may increase the utility of adenovirus vectors and also provides a means to efficiently quantify and possibly propagate HSV vectors defective in ICP0. Importantly, the results demonstrate that the activation function of ICP0 may not result from changes in cellular gene expression, but possibly as a direct consequence of an enzymatic function inherent to the protein that may involve its action at ND10 resulting in the preferential activation of viral genomes.
Palindromic Sequence Plays a Critical Role in Human Foamy Virus Dimerization
Authors: Cain Dionne , Erlwein Otto , Grigg Andrew , Russell Rebecca A. , McClure Myra O. 
Published: July 2002

ABSTRACTThe retroviral RNA genome is dimeric, consisting of two identical strands of RNA linked near their 5′ ends by a dimer linkage structure. Previously it was shown that human foamy virus (HFV) RNA transcribed in vitro contained three sites, designated SI, SII, and SIII, which contributed to the dimerization process (O. Erlwein, D. Cain, N. Fischer, A. Rethwilm, and M. O. McClure, Virology 229:251–258, 1997). To characterize these sites further, a series of mutants were designed and tested for their ability to dimerize in vitro. The primer binding site and a G tetrad in SI were dispensable for dimerization. However, a mutant that changed the 3′ end of SI migrated slower on nondenaturing gels than wild-type RNA dimers. The sequence composition of the SII palindrome, consisting of 10 nucleotides, proved to be critical for in vitro dimerization, since mutations within this sequence or replacement of the sequence with a different palindrome of equal length impaired in vitro dimerization. The length of the palindrome also seems to play an important role. A moderate extension to 12 nucleotides was tolerated, whereas an extension to 16 nucleotides or more impaired dimerization. When nucleotides flanking the palindrome were mutated in a random fashion, dimerization was unaffected. Changing the SIII sequence also led to decreased dimer formation, confirming its contribution to the dimerization process. Interesting mutants were cloned into the infectious molecular clone of HFV, HSRV-2, and were transfected into BHK-21 cells. Mutations in SII that reduced dimerization in vitro also abolished virus replication. In contrast, constructs containing mutations in SI and SIII replicated to some extent in cell culture after an initial drop in viral replication. Analysis of the SIM1 mutant revealed reversion to the wild type but with the insertion of an additional two nucleotides. Analysis of cell-free virions demonstrated that both replication-competent and replication-defective mutants packaged nucleic acid. Thus, efficient dimerization is a critical step for HFV to generate infectious virus, but HFV RNA dimerization is not a prerequisite for packaging.
Salt Separation by Charged Membranes
Authors: Schauer Jan , Kobayashi Takaomi 
Published: July 2002

Two negatively charged membranes made from poly(acrylonitrile-co-sodium styrenesulfonate) having ion exchange capacity of 0.058 and 0.15 meq/g dry matter were tested in ultrafiltration of aqueous solutions of dextrans and low-molecular-weight electrolytes. The former membrane, though permeable to bulky dextrans (molecular weight cut-off approximately 500 000), fairly retained small electrolytes. The latter, extremely hydrophilic membrane showed little resistance to the feed solution and no retention of solutes was observed. The influence of the feed concentration, electrolyte type, and operating pressure on the solute retention was determined.
Is transcutaneous electrical nerve stimulation an effective analgesia during colonoscopy?
Authors: Robinson R , Darlow S , Wright S J , Watters C , Carr I , Gadsby G , Mayberry J 
Published: July 2002

Abstract

OBJECTIVES
To evaluate the efficacy of transcutaneous electrical nerve stimulation (TENS) as analgesia during colonoscopy.

DESIGN
In a randomised controlled trial, patients undergoing diagnostic colonoscopy were assigned to one of three groups: standard medication only (midazolam); active TENS plus standard medication; or non-functioning TENS and standard medication. Efficacy of TENS was determined using numerical rating scores for pain and the post-procedural evaluation questionnaire.

SETTING
Patients undergoing diagnostic colonoscopy in a teaching hospital.

MAIN OUTCOME
There was no statistically significant differences between the three groups. However in the active TENS group there was a greater variation in “physical discomfort” and “psychological distress”, suggesting TENS may be effective in subgroup of patients.
q-SYMMETRIES IN DNLS-AL CHAINS AND EXACT SOLUTIONS OF QUANTUM DIMERS
Authors: ELLINAS DEMOSTHENES , MANIADIS PANAGIOTIS 
Published: July 2002

Dynamical symmetries of Hamiltonians quantized models of discrete nonlinear Schrödinger chain (DNLS) and of Ablowitz–Ladik chain (AL) are studied. It is shown that for n-sites the dynamical algebra of DNLS Hamilton operator is given by the su(n) algebra, while the respective symmetry for the AL case is the quantum algebra suq(n). The q-deformation of the dynamical symmetry in the AL model is due to the non-canonical oscillator-like structure of the raising and lowering operators at each site. Invariants of motions are found in terms of Casimir central elements of su(n) and suq(n) algebra generators, for the DNLS and QAL cases respectively. Utilizing the representation theory of the symmetry algebras we specialize to the n=2 quantum dimer case and formulate the eigenvalue problem of each dimer as a nonlinear (q)-spin model. Analytic investigations of the ensuing three-term nonlinear recurrence relations are carried out and the respective orthonormal and complete eigenvector bases are determined. The quantum manifestation of the classical self-trapping in the QDNLS-dimer and its absence in the QAL-dimer, is analysed by studying the asymptotic attraction and repulsion respectively, of the energy levels versus the strength of nonlinearity. Our treatment predicts for the QDNLS-dimer, a phase-transition like behaviour in the rate of change of the logarithm of eigenenergy differences, for values of the nonlinearity parameter near the classical bifurcation point.
RHEED INVESTIGATION OF Pd/Al BIMETALLIC SYSTEM ON KCl(001) SUBSTRATE
Authors: MASEK K. , MOROZ V. , MATOLÍN V. 
Published: July 2002

Pd/Al alloys have very interesting properties from the point of view of their possible application in heterogeneous catalysis. Preparation of small heteroepitaxial Pd/Al alloy particles opens a new way in studies of the influence of Pd/Al crystallographic structure on the alloy catalytic properties. Pd/Al alloy particles were grown by the molecular beam epitaxy method. Their crystallographical structure was controlled by reflection high energy electron diffraction (RHEED). It was found that Pd deposited on epitaxial 3D Al particles grown on KCl is intermixing with Al. This process is accompanied by the variation of lattice parameter from the Al value to the Pd one.
BRINGING REAL AND VIRTUAL WORLDS TOGETHER IN THE MANUFACTURING PROCESS
Authors: DIETRICH UTE , SCHULZ THOMAS , YARAMANOGLU NEZIH 
Published: July 2002

A wide spectrum of IT-tools and a variety of different systems are currently in place to provide best-in-class solutions for planning and executing manufacturing tasks. Among such applications, simulation systems play a major role in the areas of manufacturing planning and quality assurance, as well as process optimization. These systems are found in most production environments. Typically, simulation systems help to anticipate the behavior of physical systems and review the design and planning data. Today, the major automotive companies and their IT-vendors work on concepts to cover the entire process of production. However, there are still technological limitations originating from the inadequate information flow between the real and virtual worlds. A number of research projects focus on the improvement of planning processes by feeding information on life cycle conditions back to the simulation environment or bringing simulation functionality closer to manufacturing processes. The following article extracts some examples from the area of simulation systems research with a focus on the manufacturing environment and related task areas. Application cases are presented as sample scenarios for the integrated operation of virtual and real processes.
Extending relational algebra and relational calculus with set-valued attributes and aggregate functions
Authors: Özsoyoğlu G. , Özsoyoğlu Z. M. , Matos V. 
Published: July 2002

In commercial network database management systems, set-valued fields and aggregate functions are commonly supported. However, the relational database model, as defined by Codd, does not include set-valued attributes or aggregate functions. Recently, Klug extended the relational model by incorporating aggregate functions and by defining relational algebra and calculus languages.
In this paper, relational algebra and relational calculus database query languages (as defined by Klug) are extended to manipulate set-valued attributes and to utilize aggregate functions. The expressive power of the extended languages is shown to be equivalent. We extend the relational algebra with three new operators, namely, pack, unpack, and aggregation-by-template. The extended languages form a theoretical framework for statistical database query languages.