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PubMed1989

Cloning and expression in Escherichia coli of a rat hepatoma cell cDNA coding for 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase.

In liver, the 470-residue bifunctional enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFK-2/FBPase-2) catalyses the synthesis and degradation of fructose 2,6-bisphosphate, a potent stimulator of glycolysis. In rat hepatoma (HTC) cells, this enzyme has kinetic, antigenic, and regulatory properties, such as insensitivity to cyclic AMP-dependent protein kinase and lack of associated FBPase-2 activity, that differ from those in liver. To compare the sequence of the HTC enzyme with that of the liver enzyme, we have cloned the corresponding fully-coding cDNA from HTC cells. This cDNA predicts a protein of 448 residues in which the first 32 residues of liver PFK-2/FBPase-2 including the cyclic AMP target sequence have been replaced by a unique N-terminal decapeptide. The rest of the protein is identical with the liver enzyme. An N-terminally truncated recombinant peptide of 380 residues containing the PFK-2 and FBPase-2 domains was expressed in Escherichia coli as a beta-galactosidase fusion protein. It was recognized by anti-PFK-2 antibodies but its enzymic activities were barely detectable. In contrast, a cDNA fully-coding for the HTC enzyme could be expressed in E. coli as a beta-galactosidase-free peptide that exhibited both PFK-2 and FBPase-2 activities. This peptide had those PFK-2 kinetic properties of the HTC enzyme that differ from the liver enzyme. These data, together with immunoblot experiments, suggest that the lack of associated FBPase-2 activity in HTC cells results from a post-translational modification of the enzyme rather than from the difference in amino acid sequence. As well as this peculiar type of PFK-2/FBPase-2 mRNA, HTC cells also contained low concentrations of the liver-type mRNA. Unlike in liver, neither mRNA was induced by dexamethasone in these cells.

Amino Acid Sequence↗
PubMed2003

Discrete coding of reward probability and uncertainty by dopamine neurons.

Uncertainty is critical in the measure of information and in assessing the accuracy of predictions. It is determined by probability P, being maximal at P = 0.5 and decreasing at higher and lower probabilities. Using distinct stimuli to indicate the probability of reward, we found that the phasic activation of dopamine neurons varied monotonically across the full range of probabilities, supporting past claims that this response codes the discrepancy between predicted and actual reward. In contrast, a previously unobserved response covaried with uncertainty and consisted of a gradual increase in activity until the potential time of reward. The coding of uncertainty suggests a possible role for dopamine signals in attention-based learning and risk-taking behavior.

Animals↗
PubMed2003

Rats have trouble associating all three parts of the time-place-event memory code.

The ability of animals to associate an event with predictable time and place information confers a major biological advantage. The current research uses a variety of procedures and paradigms (e.g. place preference, radial arm maze, Morris water maze, T-maze, go no-go) to show that rats, unlike pigeons [e.g. Anim Learn Behav 22 (1994) 143] do not readily make an event-time-place association. They do make associations between event-time and event-place information, however. These findings are in disagreement with Gallistel's (The Organization of Learning, MIT Press, Cambridge, MA ) theory that claims that animals automatically store a memory code that has these three pieces of information. The present research is in line with the work of others who also find that rats do not readily make daily time-place associations [Behav Processes 23 (1997) 232; Behav Processes 52 (2000) 11; Behav Processes 49 (2000) 21; Anim Learn Behav 28 (2000) 298]. An interesting finding that did emerge from the present research was that at least some rats can use a circadian timer to solve a time-of-day discrimination if the task is a go no-go discrimination.

Journal Article↗
PubMed2004

Estimating the safety performance of urban road transportation networks.

Transportation planning models are typically used to estimate future traffic patterns, peak period traffic, travel time, and various environmental or other related traffic flow characteristics. Unfortunately, traffic safety is seldom, if ever, explicitly considered proactively during the transportation planning process. This omission is attributed to various factors, including the lack of available tools needed to estimate the number of crashes during this process. To help fill this void, the research on which this paper is based aimed, as a primary objective, to develop a tool that would allow the estimation of crashes on digital or coded urban transportation networks during the planning process. The secondary objective of the research was to describe how the predictive models should be applied on these networks and explain the important issues and limitations surrounding their application. To accomplish these objectives, safety performance functions specifically created for this work were applied to two sample digital networks created with the help of EMME/2, a software package widely used in transportation planning. The results showed that it is possible to predict crashes on digital transportation networks, but confirmed the reality that the accuracy of the predictions is directly related to the precision of the traffic flow estimates. The crash predictions are also sensitive to how the digital network is coded, and it is shown how appropriate adjustments can be made.

Accidents, Traffic↗
PubMed1984

Signal-to-noise ratio and bandwidth for pseudo-random codes in an ultrasonic imaging system.

Signal-to-noise ratio and image resolution in an ultrasonic imaging system are compared for four kinds of excitation signals, namely, a single pulse, an unmodulated pseudo-random code, a modulated pseudo-random code and a deconvolved pseudo-random code, all for a given length and amplitude. It was predicted and found from measurements that the best S/N and resolution are obtainable with the deconvolved pseudo-random code.

Acoustics↗
PubMed2007

Accuracy of Medicare claims data in identifying Parkinsonism cases: comparison with the Medicare current beneficiary survey.

STUDY PURPOSE: Administrative databases are commonly used to examine use of healthcare service, with researchers relying on diagnostic codes to identify medical conditions. This study evaluates the accuracy of administrative claims in identifying Parkinsonism cases compared to the self-reported Parkinson's disease (PD). METHODS: The reference cases were identified based on the self-reported PD status and the use of PD drugs collected by the 1992-2000 Medicare Current Beneficiary Survey that contained 72,922 observations from 30,469 individuals. Using ICD-9 CM, cases with PD were extracted from the corresponding Medicare claims. We compared prevalence of PD obtained using different types of claims. RESULTS: The sensitivities were the highest when all claims were used (66%). All the specificities were greater than 99%. When drug use information was included in the gold standard, the sensitivities became lower, while the specificities and positive predictive values (PPVs) increased. Using more diagnostic codes improved the sensitivity of the identification process but reduced PPVs. CONCLUSIONS: Administrative claims can provide fairly accurate and practical approach to "rule in" patients with PD. Depending on the purpose of evaluation, researchers may consider using more categories of claims to improve the sensitivity of the identification algorithm or use fewer diagnoses to minimize number of false positive cases.

Aged↗
PubMed2001

The level and landscape of optimization in the origin of the genetic code.

We consider a model of the origin of genetic code organization incorporating the biosynthetic relationships between amino acids and their physicochemical properties. We study the behavior of the genetic code in the set of codes subject both to biosynthetic constraints and to the constraint that the biosynthetic classes of amino acids must occupy only their own codon domain, as observed in the genetic code. Therefore, this set contains the smallest number of elements ever analyzed in similar studies. Under these conditions and if, as predicted by physicochemical postulates, the amino acid properties played a fundamental role in genetic code organization, it can be expected that the code must display an extremely high level of optimization. This prediction is not supported by our analysis, which indicates, for instance, a minimization percentage of only 80%. These observations can therefore be more easily explained by the coevolution theory of genetic code origin, which postulates a role that is important but not fundamental for the amino acid properties in the structuring of the code. We have also investigated the shape of the optimization landscape that might have arisen during genetic code origin. Here, too, the results seem to favor the coevolution theory because, for instance, the fact that only a few amino acid exchanges would have been sufficient to transform the genetic code (which is not a local minimum) into a much better optimized code, and that such exchanges did not actually take place, seems to suggest that, for instance, the reduction of translation errors was not the main adaptive theme structuring the genetic code.

Algorithms↗
PubMed2006

Cloning of the bovine prion-like Shadoo (SPRN) gene by comparative analysis of the predicted genomic locus.

SPRN is a new prion-like gene coding for Sho, a protein with significant similarity to PrP. SPRN was initially described in zebrafish; however, the strong evolutionary conservation led to the hypothesis that SPRN might be the ancestral prion-like gene. We mapped SPRN in Bos taurus by comparative analysis of the locus and of the predicted flanking genes. BACs, spanning the whole SPRN genomic locus, were assigned to BTA26q23 by radiation hybrid mapping and fluorescent in situ hybridization (FISH). Sequencing of five genes flanking SPRN, namely, ECHS1, PAOX, MTG1, SPRN, and CYP2E1, high-resolution FISH on mechanically stretched chromosomes, and combed BAC DNA allowed us to establish their order and reciprocal orientation. The results confirmed that BTA26q23 corresponds to HSA10q24.3-26.3, which is the site where the human SPRN is located. The gene order in Bos taurus is the same as in man, cen-ECHS1-PAOX-MTG1-SPRN-CYP2E1-tel, but PAOX has a different orientation in the two species. SPRN has the typical two-exon PRNP arrangement, with the CDS fully contained within exon 2; furthermore, it codes for a 143-amino-acid protein with 74.8% identity and 84.7% similarity with the human PRNP. RT-PCR and Northern blot analysis showed that SPRN is expressed at high levels in brain and less in testis and lung.

Animals↗
PubMed2002

Distribution of 1000 sequenced T-DNA tags in the Arabidopsis genome.

Induction of knockout mutations by T-DNA insertion mutagenesis is widely used in studies of plant gene functions. To assess the efficiency of this genetic approach, we have sequenced PCR amplified junctions of 1000 T-DNA insertions and analysed their distribution in the Arabidopsis genome. Map positions of 973 tags could be determined unequivocally, indicating that the majority of T-DNA insertions landed in chromosomal domains of high gene density. Only 4.7% of insertions were found in interspersed, centromeric, telomeric and rDNA repeats, whereas 0.6% of sequenced tags identified chromosomally integrated segments of organellar DNAs. 35.4% of T-DNAs were localized in intervals flanked by ATG and stop codons of predicted genes, showing a distribution of 62.2% in exons and 37.8% in introns. The frequency of T-DNA tags in coding and intergenic regions showed a good correlation with the predicted size distribution of these sequences in the genome. However, the frequency of T-DNA insertions in 3'- and 5'-regulatory regions of genes, corresponding to 300 bp intervals 3' downstream of stop and 5' upstream of ATG codons, was 1.7-2.3-fold higher than in any similar interval elsewhere in the genome. The additive frequency of insertions in 5'-regulatory regions and coding domains provided an estimate for the mutation rate, suggesting that 47.8% of mapped T-DNA tags induced knockout mutations in Arabidopsis.

Algorithms↗
PubMed1984

The c-sis gene encodes a precursor of the B chain of platelet-derived growth factor.

The relationship between platelet-derived growth factor (PDGF) and the proto-oncogene c-sis has been determined by amino acid sequence analysis of PDGF and nucleotide sequence analysis of c-sis genomic clones. The nucleotide sequences of five regions of the human c-sis gene which are homologous to sequences of the transforming region (v-sis) of simian sarcoma virus (SSV) were determined. By alignment of the c-sis and v-sis nucleotide sequences the predicted amino acid sequence of a polypeptide homologous to the putative transforming protein p28sis of SSV was deduced. Both predicted sequences use the same termination codon and additional coding sequences may lie 5' to the homologous regions. Amino acid sequence analysis of the PDGF B chain shows identity to the amino acid sequence predicted from the c-sis sequences over 109 amino acid residues. Polymorphism may exist at two amino acid residues. These results suggest that c-sis encodes a polypeptide precursor of the B chain. A partial amino acid sequence of the PDGF A chain is also described. This chain is 60% homologous to the B chain and cannot be encoded by that part of c-sis which has been sequenced but could be encoded by sequences which lie 5' to the five regions of v-sis homology in c-sis, or at a separate locus.

Amino Acid Sequence↗
PubMed2005

Genomic analysis of G protein gamma subunits in human and mouse - the relationship between conserved gene structure and G protein betagamma dimer formation.

Analysis of the genomic sequences, cDNAs and expressed sequence tags (ESTs) in human and mouse for the 12 genes of the gamma subunits of the heterotrimeric G proteins has allowed us to identify the common versus unique elements of the organization and expression of the members of this important gene family. All of the G protein gamma subunit genes are organized around two coding exons, each containing about 100 nucleotides coding for 30-40 amino acids. These two exons each correspond to a functional domain of the protein, which interestingly appears to impose constraints on both the structure of the protein and the structure of the gene. There is large variation in the intron size between these two coding exons, the number and size of 5' and 3' UTRs, and the overall size of the genes. There is general but not absolute conservation in the size and structure of these genes between humans and mice. Alternative splicing and potential differential promoter usage were detected for several Ggamma subunits, indicating possible differential regulation in expression. Only for Ggamma10, however, did we find an alternative coding transcript. This alternative transcript appears to code for a hybrid protein containing a DnaJ domain in place of its Ggamma exon 1 domain, joined to the Ggamma10 second exon domain. The predicted mRNA is expressed in humans, and the protein coded by it is readily translated in vitro. This protein does not form a functional G protein betagamma dimer, but it could generate a chaperone-like protein related to its DNA-J domain. These studies suggest that alternative splicing is not a prominent mechanism for generating G protein subunit diversity from within the human or mouse genomes. Instead, each of the known 12 gamma subunit genes generate transcripts with one prevalent protein.

Alternative Splicing↗
PubMed2003

The mitochondrial genome of Strongyloides stercoralis (Nematoda) - idiosyncratic gene order and evolutionary implications.

The complete mitochondrial genome sequence of the parasitic nematode Strongyloides stercoralis was determined, and its organisation and structure compared with other nematodes for which complete mitochondrial sequence data were available. The mitochondrial genome of S. stercoralis is 13,758 bp in size and contains 36 genes (all transcribed in the clockwise direction) but lacks the atp8 gene. This genome has a high T content (55.9%) and a low C content (8.3%). Corresponding to this T content, there are 16 (poly-T) tracts of >/=12 Ts distributed across the genome. In protein-coding genes, the T bias is greatest (76.4%) at the third codon position compared with the first and second codon positions. Also, the C content is higher at the first (9.3%) and second (13.4%) codon positions than at the third (2%) position. These nucleotide biases have a significant effect on predicted codon usage patterns and, hence, on amino acid compositions of the mitochondrial proteins. Interestingly, six of the 12 protein-coding genes are predicted to employ a unique initiation codon (TTT), which has not yet been reported for any other animal mitochondrial genome. The secondary structures predicted for the 22 transfer RNA (trn) genes and the two ribosomal RNA (rrn) genes are similar to those of other nematodes. In contrast, the gene arrangement in the mitochondrial genome of S. stercoralis is different from all other nematodes studied to date, revealing only a limited number of shared gene boundaries (atp6-nad2 and cox2-rrnL). Evolutionary analyses of mitochondrial nucleotide and amino acid sequence data sets for S. stercoralis and seven other nematodes demonstrate that the mitochondrial genome provides a rich source of phylogenetically informative characters. In conclusion, the S. stercoralis mitochondrial genome, with its unique gene order and characteristics, should provide a resource for comparative mitochondrial genomics and systematics studies of parasitic nematodes.

Amino Acid Sequence↗
PubMed1985

An erythromycin-resistance gene from an erythromycin-producing strain of Arthrobacter sp.

A gene (ermA) coding for a presumed erythromycin-resistance (ErR) determinant from an Er-producing Arthrobacter sp. strain (NRRLB3381) was isolated from a gene bank in phage vector lambda 2001 by probing with a Streptomyces ErR gene. Strongly hybridizing fragments were subcloned and the appropriate segments sequenced. The ermA gene is 76 mol% G + C in content and specifies a protein of 340 aa with an Mr of 37454. S1 nuclease mapping and primer extension identified the putative promoter, which resembles the consensus sequence of Escherichia coli promoters particularly in the -10 region. A potential ribosome-binding site (RBS) (AGGAG) was also located. Unexpectedly, the majority of in vivo ermA transcripts detected were only 245 nt long, suggesting that expression of ErR may be regulated post-transcriptionally. Substantial homology is observed between the predicted aa sequences of the ermA-coded protein and the products of three other ErR determinants, from organisms that do not produce Er.

Arthrobacter↗
PubMed2006

PROFtmb: a web server for predicting bacterial transmembrane beta barrel proteins.

PROFtmb predicts transmembrane beta-barrel (TMB) proteins in Gram-negative bacteria. For each query protein, PROFtmb provides both a Z-value indicating that the protein actually contains a membrane barrel, and a four-state per-residue labeling of upward- and downward-facing strands, periplasmic hairpins and extracellular loops. While most users submit individual proteins known to contain TMBs, some groups submit entire proteomes to screen for potential TMBs. Response time is about 4 min for a 500-residue protein. PROFtmb is a profile-based Hidden Markov Model (HMM) with an architecture mirroring the structure of TMBs. The per-residue accuracy on the 8-fold cross-validated testing set is 86% while whole-protein discrimination accuracy was 70 at 60% coverage. The PROFtmb web server includes all source code, training data and whole-proteome predictions from 78 Gram-negative bacterial genomes and is available freely and without registration at http://rostlab.org/services/proftmb.

Bacterial Proteins↗
PubMed1984

Cloning of human adenosine deaminase cDNA and expression in mouse cells.

A previously isolated partial cDNA sequence encoding human adenosine deaminase (ADA) was used to probe a cDNA library prepared from human cultured cell mRNA. Clones containing a combined overlapping length of 1462 bp were isolated and sequenced. One of these was found to include the entire ADA coding region. An open reading frame consisting of 363 codons was identified, predicting a polypeptide of Mr 40762. A mammalian expression plasmid was constructed, positioning the ADA coding sequence to be under transcriptional control of the mouse metallothionein promoter. Transfection of cultured mouse L-cells with this plasmid resulted in the acute expression of human ADA enzymatic activity, as assayed by isoelectric focusing.

Adenosine Deaminase↗
PubMed2001

Assessing future possible selves by gender and socioeconomic status using the anticipated life history measure.

This is a report from the first phase of a longitudinal study of the ways young adults imagine their future lives. The future possible selves of 223 18- and 19-year-old adults were examined using the Anticipated Life History measure (ALH), a psychological instrument prompting participants to describe their future life course from their 21st birthday until their death. When the ALH narratives were coded for presence/absence of life events, female participants were more likely to predict career choice, marriage, children, divorce, and death of spouse than their male counterparts; when coded for psychological qualities, female participants demonstrated greater psychological complexity and awareness of future life role choices and conflicts. Participants with lower SES wrote ALH narratives with fewer altruistic acts, less awareness of life role complexity, and fewer anticipated conflicts and their resolutions than those with higher SES.

Adolescent↗
PubMed2000

Cohesin-dockerin recognition in cellulosome assembly: experiment versus hypothesis.

The cohesin-dockerin interaction provides the basis for incorporation of the individual enzymatic subunits into the cellulosome complex. In a previous article (Pagés et al., Proteins 1997;29:517-527) we predicted that four amino acid residues of the approximately 70-residue dockerin domain would serve as recognition codes for binding to the cohesin domain. The validity of the prediction was examined by site-directed mutagenesis of the suspected residues, whereby the species-specificity of the cohesin-dockerin interaction was altered. The results support the premise that the four residues indeed play a role in biorecognition, while additional residues may also contribute to the specificity of the interaction. Proteins 2000;39:170-177.

Affinity Labels↗
PubMed1992

Molecular cloning of AMP deaminase isoform L. Sequence and bacterial expression of human AMPD2 cDNA.

Human AMPD2 cDNA clones have been isolated from T-lymphoblast and placental lambda gt11 libraries utilizing a previously cloned rat partial AMPD2 cDNA as the probe. Alignment analysis of all cDNA clones indicates the presence of intervening sequences in several placental isolates. This has been confirmed by sequencing human AMPD2 genomic clones. Intervening sequences can be removed from the cDNA clones by restriction with endonucleases at unique sites within the proposed open reading frame. This results in a 3292-base pair cDNA proposed to contain the entire AMPD2 open reading frame, which would encode a 760-amino acid polypeptide with a predicted subunit molecular mass of 88.1 kDa. Nucleotide and predicted amino acid comparisons with the 264 base pairs of proposed coding sequences in the rat AMPD2 cDNA demonstrate 91% similarity and identity, respectively. A comparison of the predicted human AMPD1 and AMPD2 polypeptides demonstrates homology in their C-terminal domains. Included in this region is the conserved motif, SLSTDDP, proposed to be part of the catalytic site of all AMP deaminases. In contrast, the predicted N-terminal domains of the human AMPD1 and AMPD2 polypeptides are unique. When placed in a prokaryotic expression vector, the human AMPD2 cDNA expresses AMP deaminase activity which can be precipitated with polyclonal antisera specific for isoform L.

AMP Deaminase↗

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