Search PubMed⌕ Search

Biomedical subjects

C Wang

Publications and source records attributed to C Wang.

At least 1,081 records · Page 60Linked to original sources
PubMed1992

Thermodynamics and structure of a DNA tetraplex: a spectroscopic and calorimetric study of the tetramolecular complexes of d(TG3T) and d(TG3T2G3T).

We report a combined thermodynamic and structural characterization of a DNA tetraplex. Using spectroscopic and calorimetric techniques, we demonstrate that d(TG3T) and d(TG3T2G3T), in the presence of K+, form stable tetramolecular complexes. From differential scanning calorimetry measurements, we obtain the following thermodynamic profiles for formation of each tetraplex at 25 degrees C: delta G degrees = -6.9 kcal/mol of tetraplex (or -2.3 kcal/mol of tetrad; 1 cal = 4.184 J), delta H degrees = -62.6 kcal/mol of tetraplex (or -20.9 kcal/mol of tetrad), and delta S degrees = -186.9 cal.K-1.mol-1 of tetraplex (or -62.3 cal.K-1.mol-1 of tetrad) for the d(TG3T) tetraplex; and delta G degrees = -20.2 kcal/mol of tetraplex (or -3.4 kcal/mol of tetrad), delta H degrees = -123.2 kcal/mol of tetraplex (or -20.5 kcal/mol of tetrad), and delta S degrees = -346.0 cal.K-1.mol-1 of tetraplex (or -57.7 cal.K-1.mol-1 of tetrad) for the d(TG3T2G3T) tetraplex. These data demonstrate that at 25 degrees C a G-tetrad can exhibit considerable stability, comparable to or even exceeding that of most Watson-Crick nearest-neighbor interactions, with this stability resulting from a very favorable enthalpy of formation. Temperature-dependent CD measurements reveal that the melting temperatures of both tetraplexes exhibit unusually low salt dependences. This unexpected behavior may reflect a diminished charge density due to bound K+ ions. For each complex, the Na+ and K+ forms exhibit drastically different isothermal and temperature-dependent CD profiles, with the K+ forms of each tetraplex melting more sharply and at a higher temperature than the Na+ forms. Using one- and two-dimensional NMR techniques, we show that the strands in the tetramolecular complex of d(TG3T), K+ are all parallel and that the guanine glycosidic conformations are all anti.

Base Sequence↗
PubMed1992

Methylations of 70,000-Da heat shock proteins in 3T3 cells: alterations by arsenite treatment, by different stages of growth and by virus transformation.

We have characterized the basic amino acid methylation of three members of the 70,000-Da heat shock protein superfamily, hsp68, hsc70, and BiP, in Balb/c 3T3 cells. It appears that a lysyl residue is the only methylation site in BiP and that both lysyl and arginyl residues are methylated in hsp68 and hsc70. In all cases, epsilon-N-trimethyllysine is the predominant methyllysine species. Both NG-monomethylarginine and NG,NG-dimethylarginine are identified as the methylarginine species. The stoichiometry of the methylation is indirectly determined by using the amount of actin methylation as a reference. Three, four, and four methyl groups are incorporated into lysyl residues of hsp68, hsc70, and BiP, respectively. The level of lysyl methylation in hsc70 remains unchanged under different growth conditions. On the other hand, the arginyl methylation in hsc70 varies considerably. In confluent Balb/c 3T3 cells, there are 1.8 and 1.3 methyl groups in dimethylarginine and monomethyl-arginine, respectively. In nonconfluent cells, the amount of monomethylarginine is similar to that in confluent cells, but dimethylarginine is not detectable. Furthermore, in both confluent and nonconfluent cells, the level of monomethylarginine is reduced 5- to 10-fold after arsenite treatment. However, in 3T3 cells transformed by Rous sarcoma virus (SR-RSV 3T3 cells), the level of arginine methylation is constitutively lower and cannot be reduced further by arsenite.

3T3 Cells↗
PubMed1992

Differential scanning calorimetry of chicken erythrocyte nuclei.

Investigation of structural features of native chromatin requires the use of intact nuclei, a turbid material which cannot be analyzed by optical methods. Differential scanning calorimetry does not require optically clear samples and has been proved by a number of authors to be a powerful tool in this field of study. By this technique, chicken erythrocyte nuclei were found to undergo at least four thermal transitions, centered at 59, 74, 88 and 98 degrees C. The highest temperature transition is strongly dependent on age and storage conditions of the nuclei. Adequate storage conditions overcame this problem and reproducible scans were obtained over a period of several months. This technical improvement has permitted the reconsideration of the occurrence of the fourth calorimetric transition, previously believed to be displayed only in replicating nuclei. Evidence gathered in the presence of perturbants and possible ligands allows the assignment of the four transitions to a nuclear protein scaffold, histones, nucleosomal DNA and a superstructured form of DNA. Moreover, it suggests that the higher-order structure is stabilized by fibronectin-like proteins.

Animals↗
PubMed1992

Characterization of human androgen receptor overexpressed in the baculovirus system.

An essential step in the process of understanding the structure and function of the human androgen receptor (hAR) involves the production of large quantities of the hAR. For this purpose, the full-length hAR has been overproduced in insect cells by using a baculovirus genetic expression system. The recombinant hAR is produced in Sf21 insect cells at approximately 7 pmol/mg of protein (1 x 10(6) AR molecules per cell), which is 70-150 times greater than levels detected in androgen target tissues. Androgen can bind to the baculovirus-expressed hAR with high affinity (Kd = 0.46 nM), and the specificity of hormone binding in baculovirus-expressed hAR is essentially identical to that of bona fide hAR. An anti-AR monoclonal antibody can recognize the baculovirus-expressed hAR at approximately 100 kDa upon Western blot analysis. Sucrose gradient analysis shows that baculovirus-expressed hAR complexes sediment at 4 S in a high salt medium and these complexes can interact with anti-AR monoclonal antibody to form complexes that sediment at 8-10 S. Therefore, production of recombinant hAR from the baculovirus expression system will provide an alternative source of biologically active hAR for studies on the molecular mechanisms of androgen action.

Androgens↗
PubMed1992

Biochemical characterization and substrate specificity of rat prostate kallikrein (S3): comparison with tissue kallikrein, tonin and T-kininogenase.

A tissue kallikrein-like enzyme encoded by S3 mRNA was purified to homogeneity from rat prostate gland. The apparent molecular mass of the prostate enzyme is 32 kDa as determined by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The intact 32 kDa enzyme is split into two bands of lower molecular mass, 18 and 14 kDa, under reducing conditions on SDS-PAGE. NH2-terminal amino acid sequence analyses of the intact enzyme and heavy and light chains revealed the identity to the translated sequence of a prostate kallikrein cDNA (S3). Isoelectric focusing indicated that the prostate enzyme is a basic protein with pI of 7.30-7.45. Specific activities of the prostate kallikrein toward angiotensin I, angiotensinogen and rat low M(r) kininogen as well as tripeptide chromogenic substrates were compared with those of tissue kallikrein, tonin and T-kininogenase. The kinin-releasing activity is inhibited by leupeptin, antipain, benzamidine and soybean trypsin inhibitor. A sensitive and specific radioimmunoassay for the rat prostate kallikrein shows that the immunoreactive kallikrein levels in prostate and submandibular gland were 23.78 +/- 2.62 micrograms/mg protein (n = 5) and 12.29 +/- 2.25 micrograms/mg protein (n = 5), respectively. The results indicate that the prostate kallikrein S3 is expressed at high levels in both prostate and submandibular glands.

Amino Acid Sequence↗
PubMed1992

Substrate specificities of tissue kallikrein and T-kininogenase: their possible role in kininogen processing.

The present studies demonstrate the importance of subsite interactions in determining the cleavage specificities of kallikrein gene family proteinases. The effect of substrate amino acid residues in positions P3-P'3 on the catalytic efficiency of tissue kallikreins (rat, pig, and horse) and T-kininogenase was studied using peptidyl-pNA and intramolecularly quenched fluorogenic peptides as substrates. Kinetic analyses show the different effects of D-amino acid residues at P3, Pro at P'2, and Arg at either P'1 or P'3 on the hydrolysis of substrates by tissue kallikreins from rat and from horse or pig. T-Kininogenase was shown to differ from tissue kallikrein in its interactions at subsites S2, S'1, and S'2. As a result of these differences, Abz-FRSR-EDDnp with Arg at P'2 is a good substrate for tissue kallikreins from horse, pig, and rat but not for T-kininogenase. Abz-FRRP-EDDnp and Abz-FRAPR-EDDnp with Pro at P'2 (rat high molecular weight kininogen sequence) are susceptible to rat tissue kallikrein but not to tissue kallikreins from horse and pig. Arg at P'3 increased the susceptibility of the Arg-Ala bond to rat tissue kallikrein. These data explain the release of bradykinin by rat tissue kallikrein and of kallidin by tissue kallikreins from other animal species. Abz-FRLV-EDDnp and Abz-FRLVR-EDDnp (T-kininogen sequence) are good substrates for T-kininogenase but not for tissue kallikrein. Arg at the leaving group (at either P'1, P'2, or P'3) lowers the Km values of T-kininogenase while Val at P'2 increases its kcat values.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1992

Structure of the rat gonadotropin releasing hormone (rGnRH) gene promoter and functional analysis in hypothalamic cells.

The gonadotropin releasing hormone (GnRH) gene encodes a protein which plays a critical role in mammalian reproductive physiology. Its expression is predominantly restricted to the hypothalamus although it has also been described in the placenta. To begin to determine the promoter elements important for tissue specific expression and to examine the mechanisms of developmental and hormonal regulation of the rat GnRH (rGnRH) gene, we cloned the rGnRH gene from a rat liver genomic DNA library. The nucleotide sequence of greater than 3 kb of 5'-flanking region was determined. The transcriptional initiation site in rat hypothalamic tissue and a mouse hypothalamic cell line were mapped by primer extension analysis and found to be different. In addition, transient transfection studies demonstrated that multiple regions of the distal promoter are important for tissue specific and basal promoter activity in hypothalamic cells. Furthermore, in these cells a potent activation region resides between -3026 and -1031 bp and suppressor region between -1031 and -903 bp upstream of the transcriptional start site. We conclude that different portions of the 5'-flanking region, which are activating and suppressing in nature, are critical for hypothalamic expression of the rGnRH gene.

Animals↗
PubMed1992

Pattern of glucose transporter (Glut 1) expression in embryonic brains is related to maturation of blood-brain barrier tightness.

A constant supply of blood-borne glucose is vital to cerebral metabolism. Although transport of glucose into the nervous tissue, effectively separated from the blood by a functional barrier (the blood-brain barrier, BBB), is one of the essential properties of the cerebral endothelium, little is known about its metabolic regulation and developmental expression in the BBB. In this study we provide evidence by immunocytochemistry that the pattern of the brain endothelial glucose transporter in rat brains (BBB-GT), immunologically homologous with the human hepatoma (G2), human erythrocyte transporter (Glut 1), changes with BBB maturation. While the neuroepithelium at embryonic days 12 and 13 shows a high incidence of immuno-detectable BBB-GT, vascularisation of the cerebral anlage and subsequent development of vascular tightness, as evidenced by intravascularly applied horseradish peroxidase and fluorescinated dextrans, is accompanied by a significant reduction of BBB-GT expression in neuroepithelial cells and confinement of BBB-GT expression to the cerebral endothelium. Immunoblots and Northern blots of embryonic brain homogenates corroborate this change in BBB-GT expression in the brain anlage at the time of BBB maturation. However, low molecular weight glucose transporters, presumed to be of non-endothelial origin, are less dramatically reduced. The development of BBB tightness, therefore, seems to play a pivotal role in the pattern of BBB-GT expression during brain differentiation.

Animals↗
PubMed1992

Effect of helium-neon laser irradiation on serum lipid peroxide concentrations in burnt mice.

The effect of helium-neon irradiation on serum lipid peroxide concentrations in mice following 6-7% body surface area burns was investigated in a controlled study. Immediately following injury by an 8-sec 100 degrees C scalding, 25 mice were irradiated by a helium-neon laser at 0.05 J/cm2. A control group of the same size underwent identical treatment but received only sham irradiation. Serum lipid peroxide concentrations increased markedly in the control group at 0.5-4 h (P less than 0.0001, two sample t-test). In the laser treated group, the lipid peroxide concentrations remained relatively constant and were significantly depressed relative to the control group 4 h following burning (P less than 0.0001, two-sample t-test).

Animals↗