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PubMed2000

The viral RNA 3'- and 5'-end structure and mRNA transcription of infectious salmon anaemia virus resemble those of influenza viruses.

The nucleotide sequences of the termini of two of the genomic segments of the negative strand RNA virus infectious salmon anaemia virus (ISAV) were determined. The sequence of the terminal 9 nucleotides at both ends of the viral RNAs was identical, and showed distinctive sequence homology with the conserved terminal sequences found in the orthomyxoviruses. For both ISAV genomic segments a computer-based secondary structure modelling indicated that the terminal 21-24 nucleotides were able to form self-complementary panhandle structures. Comparison with ISAV-derived mRNA sequences showed that ISAV mRNAs have heterogeneous 5'-ends, and are polyadenylated from a signal sequence 13-14 nucleotides downstream of the 5'-end terminus of the vRNA. Furthermore, the in vitro replication of ISAV was hindered by the RNA polymerase II inhibitor alpha-amanitin. These findings indicate that the mechanisms for replication of ISAV are similar to those of the orthomyxoviruses, and add to the previously reported structural similarities between ISAV and the orthomyxoviruses.

3' Untranslated Regions↗
PubMed1995

A mathematical model of flow through the terminal lymphatics.

Proper understanding of the mechanisms of fluid absorption and flow through the terminal lymphatics is essential for the control of several pathological conditions such as edema, bedsores and cancer. A mathematical model of the terminal lymphatics was developed using the principles of mechanics. Computer simulation results substantiate the hypothesis that fluid absorption and flow through the terminal lymphatics occur due to suction mechanisms of the adjacent contractile lymphatic segments and due to periodic fluctuations in the interstitial fluid pressure. In addition, the results suggested that increasing the length of a terminal lymphatic vessel beyond a certain limit does not cause further increase in fluid flow into the terminal lymphatic.

Biomedical Engineering↗
PubMed1996

Effects on protein structure and function of replacing tryptophan with 5-hydroxytryptophan: single-tryptophan mutants of the N-terminal domain of the bacteriophage lambda repressor.

Conformational energy computations have been carried out on the N-acetyl-N'-methylamide of 5-hydroxytryptophan (5OH-Trp) using ECEPP/3. As observed with tryptophan (Trp), the most preferred conformation about the C alpha-C beta bond of the side chain is g+ or t. This preference is reduced to only the t conformational state when 5-hydroxyTrp is in the middle of a right-handed poly(L-alanine) alpha-helix. A similar result has been obtained with Trp [Piela et al. (1987), Biopolymers 1987, 1273-1286]. These results suggest that replacement of Trp by its analog 5-hydroxyTrp may be tolerated in an alpha-helix. To test this hypothesis, we have replaced Trp by 5OH-Trp in the fifth helices of two functionally active mutants of the N-terminal domain of the bacteriophage lambda repressor. Computations on the packing of these helices have shown that no significant structural changes results from the replacement of Trp by 5OH-Trp. The DNA-binding activity of these mutants, as assessed indirectly through geometrical parameters, is also unaltered.

5-Hydroxytryptophan↗
PubMed1985

A comprehensive computer system for anesthetic record retrieval.

We have developed computer software to store data on all surgical and obstetrical anesthetics administered by our department. The computer system provides information for monitoring the residency training program, department and operating room management, professional fee billing, and research. It imposes little additional workload on our clinical personnel, who use simple codes to record the necessary data directly on the anesthetic record. Department secretarial staff transcribe data from the anesthesia and operating room records into the computer file, which is then available for producing scheduled reports and for answering inquiries from a video terminal. The system employs extensive manual and computer verification to minimize errors and omissions in the data. We report design details and more than 3.5 years experience with this system, which is now used at four affiliated teaching hospitals, has over 50,000 cases on file, and adds more than 1800 cases monthly.

Anesthesia↗
PubMed1993

The projections of intracellularly labeled auditory nerve fibers to the dorsal cochlear nucleus of cats.

The cochlear nucleus receives incoming auditory nerve discharges, preserves or transforms the signals, and distributes outgoing activity to higher centers. The organization of auditory nerve input to the cochlear nucleus will heavily influence the mechanisms by which acoustic information is processed. In order to study structure-function relationships between auditory nerve and cochlear nucleus, the axonal arborizations of type I spiral ganglion cells were labeled with intracellular injections of horseradish peroxidase after first being electrophysiologically characterized by recording with a micropipette inserted into the axon. For each auditory nerve fiber, spontaneous discharge rate (SR) and a frequency tuning curve were determined. The tuning curve yielded the characteristic frequency (CF, that frequency to which the fiber is most sensitive) and CF threshold in dB SPL. Individual axonal arborizations including all terminal swellings were reconstructed through serial sections with the aid of a light microscope and drawing tube. On average, 13.4 +/- 8.1% of the terminal swellings were found in the dorsal cochlear nucleus (DCN) and the remaining terminal swellings were located in the ventral cochlear nucleus. In the DCN, the terminal fields of auditory nerve fibers were restricted to layer III, contributed to cytoarchitectonic striations, and exhibited a systematic relationship between fiber CF and position along the strial (or long) axis of the nucleus. Computer-aided rotations revealed that the terminal fields were anisotropic, being flattened within the trans-strial axis. The maximal width of the terminal fields along the strial axis ranged from 31-321 microns and was inversely related to fiber CF and SR. Variation in the number of terminals or depth of the terminal field within layer III was not related to SR grouping or CF of the fiber.

Animals↗
PubMed1997

Light microscopic image analysis system to quantify immunoreactive terminal area apposed to nerve cells.

The present report describes a desktop computer-based method for the quantitative assessment of the area occupied by immunoreactive terminals in close apposition to nerve cells in relation to the perimeter of the cell soma. This method is based on Fast Fourier Transform (FFT) routines incorporated in NIH-Image public domain software. Pyramidal cells of layer V of the somatosensory cortex outlined by GABA immunolabeled terminals were chosen for our analysis. A Leitz Diaplan light microscope was employed for the visualization of the sections. A Sierra Scientific Model 4030 CCD camera was used to capture the images into a Macintosh Centris 650 computer. After preprocessing, filtering was performed on the power spectrum in the frequency domain produced by the FFT operation. An inverse FFT with filter procedure was employed to restore the images to the spatial domain. Pasting of the original image to the transformed one using a Boolean logic operation called 'AND'ing produced an image with the terminals enhanced. This procedure allowed the creation of a binary image using a well-defined threshold of 128. Thus, the terminal area appears in black against a white background. This methodology provides an objective means of measurement of area by counting the total number of pixels occupied by immunoreactive terminals in light microscopic sections in which the difficulties of labeling intensity, size, shape and numerical density of terminals are avoided.

Animals↗
PubMed1999

A polylinker-derived sequence, PL, highly increased translation efficiency in Escherichia coli.

Pokeweed (Phytolacca americana) antiviral protein (PAP) is a highly specific ribosome-inactivating glycosidase. The PAP gene was isolated and cloned in an expression vector containing a polylinker-derived sequence (PL) but devoid of a Shine-Dalgarno (SD) sequence. Surprisingly, E. coli cells transformed with this vector produced over twice the amount of PAP than that with the consensus SD sequence. Computer analysis of the 5' terminal region of PAP mRNA revealed a nucleotide sequence (ACCUACUCGAGUUAG) which was complementary to two domains in 16S rRNA. The heptanucleotide ACCUACU (box I) is complementary to nucleotides 1434-1440 and the GAGUUAG (box II) to nucleotides 507-513 in 16S rRNA of E. coli. To examine the role of this sequence in the translation of PAP mRNA, single or both boxes were mutated and the protein yield was measured. Mutation of box I and of box II resulted in a 2.7 and 5.3 fold decrease in protein yield respectively, indicating that the PAP gene expression was dependent on the presence of both boxes. To investigate whether PL also increases expression of other genes, human calcitonin monomeric and tetrameric genes were used as reporters. It was found that the expression level was doubled compared to that by SD. These results demonstrate that the PL is an efficient translational initiator and may be used for high level expression of certain genes in E. coli. The possible mechanisms for the high level expression are discussed.

Animals↗
PubMed1983

Reliability and validity of the Rational Behavior Inventory with a clinical population.

Administered to mental health outpatients (N = 75) who were receiving Psychotherapy the Rational Behavior Inventory as well as several other scales of psychological adjustment prior to their first therapy session. Their therapists also completed several of these scales on their clients. This procedure was repeated for a subsample upon termination of treatment. Correlations were computed among measures, and t-tests were conducted between pre- and posttreatment scores. Significant correlations were found between all measures in the expected directions, and significant pre- to post-scale differences were identified consistent with the expected effects of treatment. These results are interpreted as initial evidence for the Rational Behavior Inventory's reliability and validity with a clinical group. Future research is suggested to examine the Rational Behavior Inventory's relationship to other measures of cognitive dysfunction.

Adaptation, Psychological↗
PubMed1991

Identification of nearest-neighbor peptides in protease digests by mass spectrometry for construction of sequence-ordered tryptic maps.

Continuous-flow fast atom bombardment mass spectrometry was used for the identification of the intermediates and end products of the tryptic digest of polypeptides throughout the time-course of the reactions. Precursor/product relationships for these peptides were determined with the aid of a simple personal computer program. The C-terminal tryptic peptides were identified by performing a tryptic digest in 50% 18O-enriched buffer which resulted in labeling of all non-C-terminal peptides with 18O. This information, along with the precursor/product correlations, was used to create a sequence-ordered tryptic map of the original polypeptide. Ion intensities of intermediate hydrolysis products are compared for enzyme to substrate ratios of 1:100 and 1:1000 (w/w) over the course of the reaction. Intermediates were found to have significantly longer lifetimes when lower levels of trypsin were present.

Amino Acid Sequence↗
PubMed1992

Tombusvirus genome may encode the sixth small protein near its 3' terminus.

A short open reading frame (ORF), ORF6, potentially encoding a polypeptide (pX) of 32-69 amino acids, was revealed upon computer translation of the 3' terminal regions of tomato bushy stunt, cymbidium ringspot, cucumber necrosis and artichoke mottled crinkle tombusviruses. ORF6 has an initiating AUG codon in a favorable context and is evaluated as expressible, judging the distribution of guanosine residues within the codons. Inspection of the alignment of the four putative products encoded by ORF6 shows statistically significant sequence conservation over 11 SD above the random expectation. Secondary structure predictions based on the Garnier method demonstrate strict conservation of a loop between two beta-strands, thus suggesting functional conservation of pXs. It is suggested that pX is not involved in tombusvirus genome replication and encapsidation in cis.

Amino Acid Sequence↗
PubMed2002

Changes in electrical resistivity of swine liver after occlusion and postmortem.

The resistivity of swine liver tissue was measured in vivo, during induced ischaemia and post-mortem, so that associated changes in resistivity could be quantified. Plunge electrodes, the four-terminal method and a computer-automated measurement system were used to acquire resistivities between 10Hz and 1 MHz. Liver resistivity was measured in vivo in three animals at 11 locations. At 10 Hz, resistivity was 758 +/- 170 ohm x cm. At 1 MHz, the resistivity was 250 +/- 40 ohm x cm. The resistivity time course was measured during the first 10 min after the liver blood supply in one animal had been occluded. Resistivity increased steadily during occlusion. The change in resistivity of an excised tissue sample was measured during the first 12h after excision in one animal. Resistivity increased during the first 2h by 53% at 10 Hz and by 32% at 1 MHz. After 2h, resistivity decreased, probably owing to membrane breakdown. The resistivity data were fitted to a Cole-Cole circle, from which extracellular resistance Re, intracellular resistance Ri and cell membrane capacitance Cm were estimated. Re increased during the first 2h by 95% and then decreased, suggesting an increase in extracellular volume. Cm increased during the first 4 h by 40%, possibly owing to closure of membrane channels, and then decreased, suggesting membrane breakdown. Ri stayed constant during the initial 6h and then increased.

Animals↗
PubMed1990

Human c-myc gene contains a regulatory site similar to consensus of interferon response sequence (IRS).

Expression of c-myc proto oncogene is regulated by multiple mechanisms. Here, we report that the consensus of the regulatory region of interferon-dependent genes, GGAAAN1-3 GAAA, was found after computer search in the 5'-terminal flank of human c-myc gene in position (-76:-67). In vitro transcription of c-myc gene fragments showed that the consensus region competes with oligonucleotide GGGAAAATGAAACT for binding to specific protein(s). This oligonucleotide was shown to bind selectively the interferon-dependent positive transcription factor. Transcription of c-myc fragments lacking 5'-terminal region up to positions -101 or +71 was initiated at two sites located in the first intron. These sites did not coincide with P1 in vivo RNA cap-site. Binding of the protein factor(s) to the regulatory region of c-myc gene -76:-67 blocked the in vitro transcription initiated in the first intron.

Base Sequence↗
PubMed1987

Analysis of disulfides present in the membrane proteins of the West Nile flavivirus.

Recently the primary structure of the structural proteins of the flaviviruses West Nile (WN) virus (Castle et al., 1985; Wengler et al., 1985) and yellow fever (YF) virus (Rice et al., 1985) have been determined. As a first step in a further characterization of the organization of the structural proteins we have now studied the disulfide bridges present in the WN virus membrane proteins. All three membrane proteins, pre M, M, and E, were analyzed. The results obtained can be summarized as follows: The pre M proteins of both WN and YF virus each contain 6 cysteine residues and the position of all of these residues is strictly conserved between both viruses. The M proteins of both viruses do not contain cysteine residues. The E proteins of these viruses contain 12 cysteines and the position of all of these residues is strictly conserved between both viruses. All cysteine residues of the WN virus-derived membrane proteins are present as intramolecular disulfides. The six disulfide bridges generated from the 12 cysteine residues in the WN virus-derived E protein have been identified as follows: Cys 1-Cys 2; Cys 3-Cys 8; Cys 4-Cys 6; Cys 5-Cys 7; Cys 9-Cys 10; Cys 11-Cys 12. The analyses of the amino acid sequence conservation between the E proteins of YF and WN virus and the characterization of the disulfides have been used to develop a description of the E protein in which the molecule is assumed to be composed of the segments R1, L1, R2, L2, and R3 followed by a membrane anchor region at the carboxy-terminal region of the molecule. Computer analyses of the hydrophilicity and of the secondary structure indicate that the R1 region might contain a cluster of viral epitopes.

Amino Acid Sequence↗
PubMed1990

Brain microdialysis studies on the control of dopamine release and metabolism in vivo.

This paper studies the actions of drugs known to release dopamine from brain tissue. Most of the theoretical background to this work has been developed in experiments on slices of brain in vitro but using in vivo microdialysis we have elaborated and extended the ideas from the in vitro experiments and been able to make a direct comparison of the mechanism of action and source of dopamine released by 5 different manipulations. The mode of action of tyramine, amphetamine, veratrine, ouabain and potassium is discussed in the light of the computer model of the nerve terminal published by Justice et al. (1988). The data may lend themselves to such an interpretation, but they could be compatible with several other models.

Animals↗
PubMed2004

Comparative proteomic analysis of human whole saliva.

Human saliva performs a wide variety of biological functions that are critical for the maintenance of the oral health. Various functions include lubrication, buffering, antimicrobial protection, and the maintenance of mucosal integrity. In addition, whole saliva may be analysed for the diagnosis of human systemic diseases, since it can be readily collected and contains identifiable serum constituents. By using proteomic approach, we have established a reference proteome map of human whole saliva allowing for the resolution of greater than 200 protein spots in a single two-dimensional polyacrylamide gel. Fifty-four protein spots, comprised of 26 different proteins, were identifies using N-terminal sequencing, mass spectrometry, and/or computer matching with protein database. Ten proteins, whose levels were significantly different when bleeding had occurred in the oral cavity, were discussed in this study. These 10 proteins include alpha-1-antrypsin, apolipoprotein A-I, cystatin A, SA, SA-III, and SN, enolase I, hemoglobin beta-chain, thioredoxin peroxiredoxin B, as well as a prolactin-inducible protein. The proteomic approach identifies candidates from human whole saliva that may prove to be of diagnostic and therapeutic significance.

Adult↗
PubMed2000

Positive charge intrinsic to Arg(37)-Arg(38) is critical for dopamine inhibition of the catalytic activity of human tyrosine hydroxylase type 1.

Tyrosine hydroxylase (TH), which converts L-tyrosine to L-3, 4-dihydroxyphenylalanine, is a rate-limiting enzyme in the biosynthesis of catecholamines; its activity is regulated by the feedback inhibition of the catecholamine products including dopamine. To rationalize the significant role of the N-terminal sequence Arg(37)-Arg(38) of human TH type 1 (hTH1) in determining the efficiency of feedback inhibition, we produced mutants of which the positively charged Arg(37)-Arg(38) site was replaced by electrically neutral Gly and/or negatively charged Glu and analyzed the degree of inhibition of these mutant enzymes by dopamine. The replacement of Arg by Gly reduced the inhibitory effect of dopamine on the catalytic activity measured in the basic pH range and the replacement of Arg by Glu was enough to abolish the inhibitory effect, although these mutations brought no significant changes to the circular dichroism spectrum. The prediction of the secondary structure of N-terminal residues 1-60 by computer software specified the location of the Arg(37)-Arg(38) sequence in the turn intervening between the two alpha-helices (residues 16-29 and residues 41-59). These results suggest that the positive charge of the amino acid residues at positions 37 and 38 is one of the main factors that maintains the characteristic of the turn and is responsible for the enzyme inhibition by dopamine.

Amino Acid Sequence↗
PubMed1992

Analysis of the Escherichia coli genome: DNA sequence of the region from 84.5 to 86.5 minutes.

The DNA sequence of 91.4 kilobases of the Escherichia coli K-12 genome, spanning the region between rrnC at 84.5 minutes and rrnA at 86.5 minutes on the genetic map (85 to 87 percent on the physical map), is described. Analysis of this sequence identified 82 potential coding regions (open reading frames) covering 84 percent of the sequenced interval. The arrangement of these open reading frames, together with the consensus promoter sequences and terminator-like sequences found by computer searches, made it possible to assign them to proposed transcriptional units. More than half the open reading frames correlated with known genes or functions suggested by similarity to other sequences. Those remaining encode still unidentified proteins. The sequenced region also contains several RNA genes and two types of repeated sequence elements were found. Intergenic regions include three "gray holes," 0.6 to 0.8 kilobases, with no recognizable functions.

Bacterial Proteins↗