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Biomedical subjects

S Zhao

Publications and source records attributed to S Zhao.

At least 451 records · Page 25Linked to original sources
PubMed1995

Effect of 3'-untranslated region on expression of TNF alpha cDNA in Escherichia coli.

A construct pRL-rhTNF alpha 2, in which the 110 bp 3'-untranslated region of rhTNF alpha cDNA was deleted, was transformed in E.coli. The expression level of several positive transformants was determined. The results showed that the expression of pRL-TNF alpha 2 was stable and the biological characteristics of the protein product were the same as that of pRL-rhTNF alpha. Furthermore, the expression level of pRL-rhTNF alpha 2 was increased. It is suggested that the 3'-untranslated region may have some effects on gene expression. A TA-rich sequence, TTTATTA, contained in the 3'-untranslated region of pRL-rhTNF alpha may be involved in the inhibitory effect on gene expression.

Base Sequence↗
PubMed1994

Epitope mapping of the branched chain alpha-ketoacid dehydrogenase dihydrolipoyl transacylase (BCKD-E2) protein that reacts with sera from patients with idiopathic dilated cardiomyopathy.

Sera from 29 of 48 patients with idiopathic dilated cardiomyopathy (IDCM) and six of six patients with dilated cardiomyopathy (DCM) secondary to suspected viral myocarditis were shown to react with the branched chain alpha-ketoacid dehydrogenase (BCKD) complex mitochondrial proteins. Whereas sera from only 1 of 26 patients with ischemic heart disease showed reactivity against the BCKD complex protein, 0 of 30 sera from normal human volunteers, 0 of 64 sera from patients with lupus, and 0 of 34 sera from patients with rheumatoid arthritis showed detectable reactivity, denoting an element of specificity for the reactivity of sera from IDCM patients. The major reactivity was localized to the dihydrolipoyl transacylase (E2) component of BCKD complex. By using recombinant techniques, the immunodominant BCKD-E2 epitope recognized by sera from IDCM patients was localized to amino acid (aa) sequences 116 to 134. Each of the IDCM sera that reacted with the native BCKD complex was shown to react with the immunodominant peptide, as defined by a peptide inhibition ELISA and by an ELISA using the reactive peptide conjugated to BSA. Sera from IDCM patients that reacted with the native BCKD complex and the reactive peptide also showed inhibition of BCKD enzyme activity. The possible mechanisms for the induction of the Abs and the implications of these findings for the pathogenesis of IDCM are discussed.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1994

The STL1 gene of Saccharomyces cerevisiae is predicted to encode a sugar transporter-like protein.

A gene has been cloned from the yeast Saccharomyces cerevisiae which, on the basis of the deduced translation product, encodes a sugar transporter-like protein. This gene, STL1, was identified as an open reading frame (ORF) closely linked to the cinnamic-acid-resistance gene POF1 on chromosome IV. The putative translation product of STL1 (STL1) contains 536 amino acids, with a M(r) of 60,507. Hydropathy analysis of STL1 suggests that it contains the twelve transmembrane (TM) domains characteristic of a family of sugar transporters from S. cerevisiae and other organisms. STL1 displays greatest homology (28% identity) to the products of the yeast HXT2 (hexose transporter) and GAL2 (galactose transporter) genes. Disruption of STL1 had no detectable effect on yeast growth on glucose, galactose, mannose, maltose or glycerol as sole carbon source. The transport function of the gene product remains unknown at present.

Amino Acid Sequence↗
PubMed1994

A mutant of protein phosphatase-1 that exhibits altered toxin sensitivity.

A chimeric mutant was constructed in which a 4-amino acid region (GEFD, residues 274-277) of rabbit muscle protein phosphatase-1 was replaced with the sequence YRCG corresponding to residues 267-270 of rabbit protein phosphatase-2A. This was based on the findings of a gene mutation in okadaic acid-resistant cells which results in a Cys-->Gly conversion in protein phosphatase-2A. The YRCG mutant of protein phosphatase-1 was expressed and purified. The properties of the mutant enzyme were investigated in terms of its sensitivity toward several toxin inhibitors (okadaic acid, microcystin, nodularin, calyculin A, and cantharidic acid), as well as inhibitor-2. The mutant enzyme exhibited a gain of function in the form of a 10-fold increased sensitivity toward okadaic acid that suggests this region is involved in toxin binding. Significant changes in sensitivity to inhibitor-2 and several of the other toxins were also observed, indicating that these may have a common binding region.

Amino Acid Sequence↗
PubMed1994

Characterization of deletion mutants of the catalytic subunit of protein phosphatase-1.

Deletion mutagenesis was used to define the core region of the catalytic subunit of rabbit muscle protein phosphatase-1. Deletions in the N terminus were found to lead to loss of expression. Deletions of up to 33 residues from the C-terminal region were tolerated, and the truncated enzymes were fully active. Deletion of an additional 21 residues led to loss of expression. Mutants which had had 33 and 25 residues deleted maintained specific activities that were comparable to those of the wild type enzyme. The response of these two deletion mutants to okadaic acid, microcystin, and inhibitor-2 was determined. Only slightly lower IC50 values were observed in all cases, showing that the C terminus itself does not play a major role in the binding of these inhibitors. The deletion mutants formed stable complexes with inhibitor-2 as shown by gel filtration. These studies provide unambiguous evidence that the extreme C-terminal region of protein phosphatase-1 is not directly involved in catalytic function or in the binding of inhibitor-2, microcystin, or okadaic acid, and they also establish that the first approximately 300 residues of the sequence constitute a sufficient core for protein phosphatase-1 catalytic functions.

Amino Acid Sequence↗
PubMed1994

G1/S-regulated E2F-containing protein complexes bind to the mouse thymidine kinase gene promoter.

By performing DNase I footprint analysis, we had identified three distinct protein binding sequences (MT1, MT2, and MT3) located on the mouse thymidine kinase (TK) upstream promoter (Dou, Q.-P., Fridovich-Keil, J. L., and Pardee, A.B. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 1157-1161). Here we report that MT2 includes an E2F-like binding site (GTTCGCGGGCAAA), as shown by the following evidence. (i) MT2 bound specifically to an affinity-purified fusion human E2F protein. (ii) Both MT2 and an authentic E2F site (TTTCGCGCGCTTT) bound specifically to similar or identical nuclear protein complexes. (iii) Formation of both these DNA-protein complexes were cell cycle-dependent: a G0/G1 phase-specific complex (E2F.G0/G1) was replaced by an S phase-specific complex(es) (E2F.S), whereas "free" E2F increased after the G1/S transition. (iv) Pulse inhibition of protein synthesis with cycloheximide interchanged these complexes with similar kinetics. (v) When MT2-shifted E2F.G0/G1, E2F.S, and free E2F were eluted and analyzed by Western blot assay using a specific antiserum to human E2F-1, two forms of murine E2F (62 and 66 kDa) were observed from all three complexes. The compositions of these MT2-bound complexes were also investigated. Studies using specific antibodies revealed that p107, a retinoblastoma-like protein, was present in both E2F-G0/G1 and E2F.S, whereas cyclin E.cyclin A.cdk2 were only present in E2F.S complex(es). These data suggest that removal of the p107-containing E2F.G0/G1 complex, a candidate repressor, from the MT2 site in late G1 may be essential for S phase-dependent transcription of the mouse TK gene.

Animals↗
PubMed1994

Lifetime and 12-month prevalence of DSM-III-R psychiatric disorders in the United States. Results from the National Comorbidity Survey.

BACKGROUND: This study presents estimates of lifetime and 12-month prevalence of 14 DSM-III-R psychiatric disorders from the National Comorbidity Survey, the first survey to administer a structured psychiatric interview to a national probability sample in the United States. METHODS: The DSM-III-R psychiatric disorders among persons aged 15 to 54 years in the noninstitutionalized civilian population of the United States were assessed with data collected by lay interviewers using a revised version of the Composite International Diagnostic Interview. RESULTS: Nearly 50% of respondents reported at least one lifetime disorder, and close to 30% reported at least one 12-month disorder. The most common disorders were major depressive episode, alcohol dependence, social phobia, and simple phobia. More than half of all lifetime disorders occurred in the 14% of the population who had a history of three or more comorbid disorders. These highly comorbid people also included the vast majority of people with severe disorders. Less than 40% of those with a lifetime disorder had ever received professional treatment, and less than 20% of those with a recent disorder had been in treatment during the past 12 months. Consistent with previous risk factor research, it was found that women had elevated rates of affective disorders and anxiety disorders, that men had elevated rates of substance use disorders and antisocial personality disorder, and that most disorders declined with age and with higher socioeconomic status. CONCLUSIONS: The prevalence of psychiatric disorders is greater than previously thought to be the case. Furthermore, this morbidity is more highly concentrated than previously recognized in roughly one sixth of the population who have a history of three or more comorbid disorders. This suggests that the causes and consequences of high comorbidity should be the focus of research attention. The majority of people with psychiatric disorders fail to obtain professional treatment. Even among people with a lifetime history of three or more comorbid disorders, the proportion who ever obtain specialty sector mental health treatment is less than 50%. These results argue for the importance of more outreach and more research on barriers to professional help-seeking.

Adolescent↗
PubMed1994

DSM-III-R generalized anxiety disorder in the National Comorbidity Survey.

BACKGROUND: Nationally representative general population data are presented on the current, 12-month, and lifetime prevalence of DSM-III-R generalized anxiety disorder (GAD) as well as on risk factors, comorbidity, and related impairments. METHODS: The data are from the National Comorbidity Survey, a large general population survey of persons aged 15 to 54 years in the noninstitutionalized civilian population of the United States. DSM-III-R GAD was assessed by lay interviewers using a revised version of the Composite International Diagnostic Interview. RESULTS: Generalized anxiety disorder was found to be a relatively rare current disorder with a current prevalence of 1.6% but was found to be a more frequent lifetime disorder affecting 5.1% of the US population aged 15 to 45 years. Generalized anxiety disorder was twice as common among women as among men. Multivariate logistic regression analysis showed that being older than 24 years, separated, widowed, divorced, unemployed, and a homemaker are significant correlates of GAD. Consistent with studies in treatment samples, we found that GAD was frequently associated with a wide spectrum of other mental disorders, with a lifetime comorbidity among 90.4% of the people who had a history of GAD. CONCLUSION: Contrary to the traditional view that GAD is a mild disorder, we found that the majority of people with GAD, whether they were comorbid or not, reported substantial interference with their life, a high degree of professional help seeking, and a high use of medication because of their GAD symptoms. Although lifetime GAD is highly comorbid, the proportion of current GAD that is not accompanied by any other current diagnosis is high enough to indicate that GAD should be considered an independent disorder rather than exclusively a residual or prodrome of other disorders.

Adolescent↗
PubMed1994

Expression of recombinant inhibitor-2 in E. coli and its utilization for the affinity chromatography of protein phosphatase-1.

The coding sequence for rabbit muscle protein phosphatase inhibitor-2 was inserted into the pET3a expression vector. This vector allowed the expression of recombinant inhibitor-2 at levels of ca. 1.5% of the soluble protein. A simple procedure allowed the purification of inhibitor-2 from Escherichia coli lysates. This involved heat-treatment, followed by chromatography on Blue-Sepharose and Q-Sepharose. Recombinant inhibitor-2 inhibited rabbit muscle protein phosphatase-1 with a potency similar to that reported for the wild type protein. The recombinant protein was coupled to CH-Sepharose and this support was found to bind the catalytic subunit of protein phosphatase-1 with high efficiency. A procedure for a single-step affinity purification of recombinant ppase-1 from E. coli lysates was shown to be feasible.

Amino Acid Sequence↗
PubMed1994

rDlx, a novel distal-less-like homeoprotein is expressed in developing cartilages and discrete neuronal tissues.

From a rat chondrosarcoma we isolated a cDNA that encodes a novel homeoprotein rDlx. The homeodomain of rDlx shows a high degree of sequence identity with those of Drosophila Distal-less, mouse Dlx, and Xenopus Xdll proteins. Northern hybridization of rDlx revealed a 1.4- to 1.6-kb RNA species in a rat chondrosarcoma and a cell line derived from this tumor and in mouse C3H10T1/2 cells, but no rDlx RNA was detected in mouse NIH3T3 fibroblasts, rat skin fibroblasts, mouse C2 myoblasts, mouse myeloma S194 cells, human B-cell lymphoma Daudi cells, or human acute myelocytic leukemia cells. RNase protection assays showed that rDlx transcripts were present at high levels in 14-day-old rat embryos, 18-day-old rat embryo skeletal tissues, and adult rat brain. rDlx RNAs were present at lower levels in newborn rat rib cartilage, 18-day-old rat embryo soft tissues, newborn rat skin, and adult rat heart. rDlx transcripts were not detected in adult rat liver, spleen, lung, kidney, testis, or skeletal muscle. In situ hybridization of rat embryos at different stages revealed that rDlx transcripts were present in otic vesicle, branchial arches, apical ectodermal ridge of limb bud, developing cartilages, perichondria of mature cartilages, mesenchymal cells of developing membranous bones, developing teeth, ganglionic eminence of the telencephalon, diencephalon, olfactory epithelia, and epidermis of the skin. rDlx RNAs were also detected in the developing parasympathetic mesenteric ganglia of the gastrointestinal tract. Hence, rDlx RNAs are mainly expressed in several neuronal tissues and developing skeletal tissues.

Amino Acid Sequence↗
PubMed1994

Cloning, sequencing, and structural analysis of the DNA encoding inosine monophosphate dehydrogenase (EC 1.1.1.205) from Tritrichomonas foetus.

The inosine monophosphate dehydrogenase (IMPDH) of the parasitic protozoan Tritrichomonas foetus is a purine salvage enzyme with a subunit molecular weight of 58,000. The enzyme has been purified to homogeneity by Verham et al. (Molecular and Biochemical Parasitology 24, 1-12, 1987) and characterized in more detail by Hedstrom and Wang (Biochemistry 29, 849-854, 1990). We used a polyclonal antibody directed against the purified enzyme to identify three cDNA clones from T. foetus. These clones were sequenced and found to contain an open reading frame encoding 497 amino acids. By complementation studies on an Escherichia coli mutant with its IMPDH gene deleted, the cDNA clones were able to transform the bacterial cells to grow on minimal medium without guanine. One of the cDNA clones, 2aa1, was used to identify two genomic clones, 2d1c and 3m4b, both containing a 4.1-kb HindIII fragment. The fragment was subcloned into the Bluescript KS+ plasmid, sequenced, and found to contain the same open reading frame as the cDNA clone except that it encodes six additional amino acid residues at the N-terminus. Its sequence has a 34% identity with that of the human IMPDH, 32% with that of E. coli IMPDH, and 31% with that of Leishmania donovani IMPDH. The molecular weight of the deduced protein is 55,534. Two segments of polypeptide that are conserved in all other IMPDHs, containing the putative NAD+ and IMP binding sites, are also relatively conserved in T. foetus. Since the parasite enzyme differs from the bacterial and mammalian IMPDHs by a very high Km value for NAD+ and an even higher KI value for mycophenolic acid (MPA) (Verham et al. 1987; Hedstrom and Wang 1990), the sequence of the parasite enzyme may provide information on the mechanism of MPA binding and the chance for other specific inhibitor design.

Amino Acid Sequence↗
PubMed1994

Optical imaging of human breast cancer.

Since an increasing number of breast cancers have been reported in recent years, there is a need for improving techniques for early detection of the breast cancer. Here we tested a time gated optical imaging technique as a tool for imaging human breast. Pulsed laser light at wavelengths of 780 and 830 nm are transmitted through human breast tissues and time spectra of the diffused light through the tissue are recorded over nanoseconds. Data from different locations are acquired and used to reconstruct a two dimensional image as a set of spectra in pixel form. The imaging consists of absorption and scattering coefficients, and the absorption coefficients at the two wavelengths are related to oxygen concentration and blood volume. The analysis of these coefficients is based upon the early arrival photons, therefore allowing construction of a better image than those from the current diaphanography. We demonstrate images of breast cancer, cysts created after lumpectomy, and consequences of radiation therapy. Results show that time gated optical imaging can image oxygen concentration in the cancerous and fibrotic breasts. Resolution of the imaging for smaller tumor size needs to be improved.

Adult↗
PubMed1994

Analysis of protein binding to receptor-doped lipid monolayers by Monte Carlo simulation.

This paper presents a Monte Carlo simulation (MCS) method for estimating the parameters that characterize ligand-receptor binding directly from experimentally derived binding isotherms. Binding parameters are estimated by incorporating an MCS algorithm for ligand binding to a two-dimensional receptor array into a nonlinear regression program. The MCS method was tested by analyzing experimental isotherms of avidin binding to biotinylated lipid in Langmuir-Blodgett (LB) monolayers. The MCS-derived cooperativity coefficients and intrinsic association constants for avidin-biotin binding to LB films are correlated strongly (R2 > 0.93) with the binding parameters determined from the same experimental data by a thermodynamic equilibrium binding model (Zhao et al. 1993. Langmuir. 9:3166-3173). This result shows MCS to be an accurate and potentially more versatile method for characterizing biomolecular interactions at surfaces.

Algorithms↗
PubMed1994

Changes in cytochromes P-450, 2E1, 2B1, and 4A, and phospholipases A and C in the intragastric feeding rat model for alcoholic liver disease: relationship to dietary fats and pathologic liver injury.

The influence of dietary fat and alcohol on hepatic microsomal levels of cytochromes P-450 2E1, 2B, and 4A; phospholipases A and C; and UDP-glucuronosyltransferase was studied in the intragastric feeding rat model for alcoholic liver injury. Eight groups of animals were evaluated. Control and ethanol fed rats received either saturated fat or corn oil and were killed after 2 weeks and 1 month of feeding. All animals were pair-fed by continuous infusion of liquid diet through permanently implanted gastric cannulas. Alcoholic liver injury developed only in the corn oil-ethanol-fed groups and was manifest by 1 month. Livers were subjected to the following analyses: pathologic evaluation of liver injury; levels of cytochromes P-450 2E1, 2B, and 4A protein and mRNA; aniline hydroxylase activity; and phospholipase A and C and UDP-glucuronosyltransferase activities. Ethanol-induced increases in cytochromes P-450 2E1 and 2B protein determined by Western blotting were greatest in the corn oil-ethanol-fed group, which developed pathologic changes in the liver. Cytochromes P-450 2E1 and 2B1 mRNA levels were unaffected, suggesting that posttranscriptional mechanisms are responsible for the increase in the corresponding P-450 proteins. In contrast, cytochrome P-450 4A levels were higher in the saturated fat-ethanol groups compared with the corn oil-ethanol groups. Phospholipase A and phospholipase C levels were higher in the corn oil-ethanol groups compared with pair-fed dextrose controls and the saturated fat-ethanol groups. UDP-glucuronosyltransferase levels declined with time in the ethanol-fed groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Markedly enhanced cytochrome P450 2E1 induction and lipid peroxidation is associated with severe liver injury in fish oil-ethanol-fed rats.

We evaluated the role of changes in cytochrome P-450 2E1 (CYP 2E1) and lipid peroxidation in relation to development of severe liver injury in fish oil-ethanol-fed rats. The experimental animals (male Wistar rats) were divided into 5 rats/group and were fed the following diets for 1 month: corn oil and ethanol (CO+E) or corn oil and dextrose (CO+D), and fish oil and ethanol (FO+E) or fish oil and dextrose (FO+D). For each animal, microsomal analysis of CYP 2E1 protein, aniline hydroxylase activity, fatty acid composition, and conjugated dienes was conducted. Also, evaluation of severity of pathology was done for each rat. The mean +/- SD of the pathology score was significantly higher (p < 0.01) in the FO+E (6.0 +/- 1.3) group than in the CO+E group (3.0 +/- 0.5). No pathological changes were evident in the dextrose-fed controls. The CYP 2E1 protein levels (mean +/- SD) were significantly higher (p < 0.01) in the FO+E group (13.1 +/- 2.0) compared with the CO+E (4.7 +/- 1.2) and FO+D (1.8 +/- 0.5) groups. Higher levels of eicosapentaenoic and docosahexaenoic acids and lower levels of arachidonic acid were detected in liver microsomes from rats fed fish oil compared with corn oil. A significant correlation was obtained between CYP 2E1 protein and conjugated diene levels (r = 0.78, p < 0.01). Our results showing markedly increased CYP 2E1 induction and lipid peroxidation in the FO+E group provides one possible explanation for the greater severity of liver injury in this group.

Animals↗
PubMed1994

The reductive acetyl coenzyme A pathway: sequence and heterologous expression of active methyltetrahydrofolate:corrinoid/iron-sulfur protein methyltransferase from Clostridium thermoaceticum.

The methyltransferase (MeTr) from Clostridium thermoaceticum transfers the N5-methyl group of (6S)-methyltetrahydrofolate to the cobalt center of a corrinoid/iron-sulfur protein in the acetyl coenzyme A pathway. MeTr was purified to homogeneity and shown to lack metals. The acsE gene encoding MeTr was sequenced and actively expressed in Escherichia coli at a level of 9% of cell protein. Regions in the sequence of MeTr and the E. coli cobalamin-dependent methionine synthase were found to share significant homology, suggesting that they may represent tetrahydrofolate-binding domains.

5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
PubMed1994

Granulocyte-colony stimulating factor, granulocyte-macrophage colony stimulating factor, PIXY-321, stem cell factor, interleukin-3, and interleukin-7: receptor binding and effects on clonogenic proliferation in acute lymphoblastic leukemia.

Cytokines are frequently used after chemotherapy of leukemias and solid tumors to augment recovery of normal hematopoiesis. While the regulation of normal and leukemic myelopoiesis is well investigated, little is known about effects of cytokines on growth and differentiation of lymphoblastic leukemia. In this study, we investigated the expression of receptors for G-CSF, GM-CSF, SCF, IL-3, and IL-7 on acute lymphoblastic leukemia (ALL) blasts and the effects of these growth factors (GF) on ALL blast colony formation. The binding of fluorescence-tagged cytokines to receptors on ALL blasts was studied by flow-cytometry in 27 cases of ALL (24 precursor B-ALL, 3 T-ALL). Receptor-binding for myeloid-associated GF was observed in the majority of precursor B-ALL (G-CSF = 100%, GM-CSF = 65%, IL-3 = 83%, SCF = 74%), but not in T-ALL. Binding of labelled IL-7 was detected in both precursor B- (92%) and T-ALL (100%). The presence of receptors for SCF in ALL was confirmed by polymerase chain reaction for c-kit mRNA in 19/21 cases tested. Expression of receptors for G-CSF, GM-CSF, IL-3, and SCF was not associated with expression of myeloid antigens, or with specific cytogenetic abnormalities. The effects of these GF on clonogenic cells were tested in the ALL blast colony assay and varied between samples, but all cytokines were able to increase clonogenic growth. The GM-CSF/IL-3 fusion molecule PIXY-321 was most effective in promoting colony growth. In some cases inhibition of colony formation was found. We conclude that ALL blast cells have receptors not only for IL-7, but also for G-CSF, GM-CSF, SCF, and IL-3. ALL precursors can respond to these GF with changes in their clonogenic growth indicating the presence of functional receptors. Results may have implications for therapeutic approaches combining cytokines and chemotherapy.

Adolescent↗