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Biomedical subjects

T Y Liu

Publications and source records attributed to T Y Liu.

At least 109 records · Page 6Linked to original sources
PubMed1989

Identification of the cysteine residue of beta-tubulin alkylated by the antimitotic agent 2,4-dichlorobenzyl thiocyanate, facilitated by separation of the protein subunits of tubulin by hydrophobic column chromatography.

The mechanism of action of the antimitotic drug 2,4-dichlorobenzyl thiocyanate (DCBT) has been examined in detail. Shown in previous studies to inhibit tubulin polymerization [Abraham, I., Dion, R. L., Duanmu, C., Gottesman, M. M., & Hamel, E. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 6839-6843] and to form a covalent bond preferentially with beta-tubulin [Bai, R., Duanmu, C., & Hamel, E. (1989) Biochim. Biophys. Acta 994, 12-20], DCBT has now been documented to interact at low concentrations with a high degree of specificity at cysteine residue 239 of beta-tubulin. These low DCBT concentrations also result in the partial inhibition of tubulin polymerization. Such findings strongly indicate that cysteine-239 of beta-tubulin is essential for microtubule assembly. Although alpha-tubulin is alkylated almost as well as beta-tubulin when the drug:tubulin ratio = 5:1 (Bai et al., 1989), beta-tubulin is alkylated about 25 times as extensively as alpha-tubulin, almost exclusively at Cys-239, when the drug:tubulin ratio = 1:5. In addition, we find that low concentrations of DCBT do not affect the binding of colchicine to tubulin but that colchicine and related compounds do reduce the alkylation of tubulin by DCBT. This suggests that Cys-239 of beta-tubulin is not involved in the binding of colchicine to tubulin but that this amino acid residue is at least partially masked by the drug when it is bound to the protein. We also describe a column chromatography procedure (hydrophobic chromatography on decylagarose) useful for the preparative resolution of unalkylated, although denatured, alpha- and beta-tubulin.

Alkylation↗
PubMed1989

Rubella virus: mechanism of attenuation in the vaccine strain (HPV77).

The vaccine type (HPV77 strain) of rubella virus replicates slower and manifests a delayed appearance of cytopathic effect in Vero-76 cells as compared to wild-type virus (M33). The change in cytopathic effect coincides with the delayed appearance of both genomic and subgenomic RNA as well as viral structural proteins in the cell. The delay in the appearance of the viral proteins in the cells was also evident when the cells infected with the vaccine-type virus were treated with the lysosomotropic agent such as chloroquine. Binding studies using [35S]methionine-labeled virus showed that the vaccine-type virus bound to the cells poorly and the binding was not completely competed out with the cold virus.

Animals↗
PubMed1989

Steroid hormone receptors in meningiomas of Chinese patients.

Meningioma is a hormone-related intracranial tumor. Brain tumor samples were collected from 32 Chinese patients, 20 of whom had meningiomas. Estrogen receptors were present in 6 of 17 meningiomas, 1 hemangioblastoma, 1 gemistocytic astrocytoma, and 1 neurilemoma. Of the 26 samples assayed for progesterone receptors, measurable bindings were found in 14 of 17 meningiomas, 1 of 2 astrocytomas, 1 malignant lymphoma, 1 hemangioblastoma, 1 ganglioglioma, 1 glioblastoma multiforme, 1 anaplastic ependymoma, 1 small cell sarcoma, and 1 neurilemoma. Androgen receptors were found in 8 of 19 meningiomas, but not in the 11 other brain tumors examined. Measurable glucocorticoid receptors were found in 15 of 19 meningiomas, 1 malignant lymphoma, 1 hemangioblastoma, 2 craniopharyngiomas, 1 neurilemoma, 1 of 2 glioblastomas, and 1 of 2 astrocytomas. Of the 10 samples examined for tyrosine aminotransferase activity, measurable enzyme activity was found in 7 of the 8 meningiomas, but not in the single hemangioblastoma nor in the single craniopharyngioma. In accordance with other published studies, the results from this study also suggest that the growth of meningiomas may be regulated by hormones, and that the role steroid hormones and their receptors play in the meningioma is worthy of further investigation.

Adult↗
PubMed1989

Studies on peptides. CLXVII. Solid-phase syntheses and immunological properties of fragment peptides related to human malaria circumsporozoite protein.

A glycine-linked tetramer of Asn-Ala-Asn-Pro, a tandem repeated sequence of malaria circumsporozoite (CS) protein, was synthesized by the Boc-based solid phase method, followed by deprotection with 1 M trimethylsilyl trifluoromethanesulfonate-thioanisole in trifluoroacetic acid. In addition, three tetramer-related peptides were similarly synthesized, i.e., a 34-residue peptide [linked with TH, a proposed T-cell epitope of CS, at the C-terminus of the tetramer], a 46-residue peptide and a 59-residue peptide [linked with HA or HA', two proposed T-cell epitopes of influenza hemagglutinin protein, at the N-terminus of the above 34-residue peptide]. Their immunological properties were examined by enzyme-linked immunosorbent assay, for which three different congenic strains of mouse were used to raise the specific antibodies. Despite conjugation of T-cell epitopes to the tetramer, the mice of low-responder strains to the tetramer failed to produce any antibody specific to the tetramer. However, with the aid of recombinant interleukin 2 as an adjuvant, the low-responder mice produced antibody with relatively high titers.

Antigens, Surface↗
PubMed1989

Nucleotide sequence of bovine prochymosin cDNA and the mechanism of cDNA deletion.

A plasmid pTLC23 harbouring entire cDNA sequence coding for prochymosin was constructed from clones pCT5 and pCT15. The nucleotide sequence of the complete cDNA insert, determined by using dideoxy sequencing method, indicates that the cloned DNA apparently represents the coding sequence for prochymosin B. Based on the data presented here, the differences between the DNA sequences reported by Nishimori et al., Harris et al. and Moir et al. were discussed as well. DNA sequence analysis reveals that the deletion in pCT15 covers positions 862-960. The missing sequence is characterized as an entire exon between intron G and intron H of the prochymosin gene, and by ending with splicing signal AG, it is suggested that the deletion in middle region of coding sequence results from the incorrect splicing of pre-mRNA.

Amino Acid Sequence↗
PubMed1989

Analysis of monoamines in the cerebrospinal fluid of Chinese patients with neurological diseases.

Levels of dopamine (DA), 3-methoxy-4-hydroxy-phenylacetic acid (HVA), 5-hydroxy-indoleacetic acid (HIAA), 5-hydroxytryptamine (HT), 5-hydroxytryptophan (HTP), and 3, 4-dihydroxyphenylalanine (DOPA) in cerebrospinal fluids (CSF) of twenty-four Chinese with or without neurological diseases were determined simultaneously. The level of HIAA in the CSF of myelopathy-spondylosis patients and the level of DA in CSF of the myelopathy-tumor patients were higher than that of the control. No difference in levels of monoamines and their metabolites was observed between the control and patients with dementia.

Adolescent↗
PubMed1988

Glucocorticoid receptors in hepatocellular carcinoma and adjacent liver tissue.

Glucocorticoid and progesterone receptors, tyrosine aminotransferase, gamma-glutamyltransferase and alpha-fetoprotein levels were determined in human hepatocellular carcinoma (HCC) and adjacent liver tissues. Glucocorticoid receptor was present in seven of ten HCC samples, values ranged from 1.9 to 66.8 fmol/mg protein. Progesterone receptor was present in two of ten HCC samples with values of 1.7 and 7.2 fmol/mg protein, respectively. In the adjacent liver tissues, no measurable progesterone receptor was found and only one sample had glucocorticoid receptor with a value of 3.0 fmol/mg protein. The increase of glucocorticoid receptor in HCC samples was coincident with a decreased level of tyrosine aminotransferase and an increased level of gamma-glutamyltransferase. No correlation was found among glucocorticoid receptor level, serum or tissue alpha-fetoprotein levels. The presence of glucocorticoid receptors in HCC suggest that hormones may play an important role in the formation of hepatoma, and hormonal therapy may be useful for patients with HCC.

Carcinoma, Hepatocellular↗
PubMed1988

Reproducible high yield sequencing of proteins electrophoretically separated and transferred to an inert support.

A method allowing initial sequencing yields of 60-85% to be consistently obtained from samples prepared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electrophoretic transfer is described in detail. Conducting electrophoresis at a pH near neutrality is the single most important of the modifications made to earlier procedures, but pre-electrophoresis in the presence of glutathione or sodium thioglycolate and use of Immobilon polyvinylidene difluoride membranes all contribute to the success of the technique. When tryptophan was the NH2 terminus of a protein, the phenylthiohydantoin (PTH)-derivative recovered appeared to be an irreversible oxidation product if pre-electrophoresis was not performed. Following pre-electrophoresis, the PTH-derivative recovered co-migrated with that of unmodified tryptophan, and the recovery was higher. Recovery of methionine as its PTH-derivative was not affected by pre-electrophoresis suggesting that thioglycolate in the electrophoresis buffer during sample separation prevented or reversed oxidation of methionine sulfur but did not protect tryptophan.

Amino Acid Sequence↗
PubMed1988

Rubella virus replication: effect of interferons and actinomycin D.

The effect of alpha and gamma interferon (IFN alpha, IFN gamma) and actinomycin D on the expression of wild type rubella virus in African green monkey kidney cells (Vero 76) was studied. Viral protein synthesis in the infected cells was significantly reduced upon treatment of the cells with IFN alpha or IFN gamma, which is accompanied by the reduction in the level of both the (+) stranded and the (-) stranded viral RNAs. The residual rubella viral RNA from interferon-treated cells, however, was structurally intact as judged by Northern blot analysis and in vitro translation. These results suggest that the effect of IFN alpha and IFN gamma on rubella viral protein synthesis is both at the transcriptional and the translational level. The effect of actinomycin D on rubella virus replication was found to be time-dependent. It is much more pronounced during the eclipse phase of the viral growth (first 4 h) than after 8 h at which time actinomycin D had lesser effect. A similar effect on rubella virus replication was observed when alpha-amanitin was used instead of actinomycin D. These results were taken to indicate that during the viral infection, host cell DNA directs the synthesis of a cellular factor(s) which is essential for the viral replication. When the synthesis of this cellular factor(s) is terminated at an early stage of viral infection by actinomycin D or by alpha-amanitin, viral replication is impaired.

Amanitins↗
PubMed1988

Structural and functional properties of a 69-kilodalton outer membrane protein of Bordetella pertussis.

A-69-kDa outer membrane protein present on virulent Bordetella pertussis cells is recognized by the agglutinating monoclonal antibodies BPE3, BPD8, and BPE8. The amino acid composition of this protein, purified from heat extracts of B. pertussis BP353 cells, is different from that of the two major fimbrial antigens of B. pertussis, which is consistent with its being a nonfimbrial protein based on other criteria. Western blot analysis using the monoclonal antibody BPE3 demonstrated that a slightly larger but antigenically cross-reactive protein is also expressed by Bordetella bronchiseptica and Bordetella parapertussis. In addition, a large molecular weight species of about 180-kDa is found in outer membrane extracts of B. bronchiseptica which may represent a precursor form of the protein or indicate that the protein can exist as an oligomer. The monoclonal antibody BPD8 directed against the 69-kDa protein almost completely inhibited the enzymatic activity of adenylate cyclase purified from B. pertussis and also inhibited the intoxication of mammalian cells by this enzyme. Since little enzymatic activity was found associated with the purified 69-kDa protein, these data suggest a role for the 69-kDa protein in regulating the adenylate cyclase toxin of B. pertussis. An additional monoclonal antibody directed against the 69-kDa protein, BPE8, decreases lymphocytosis and delays death in mice receiving a respiratory challenge of virulent B. pertussis cells. These studies suggest that further investigation into the role of this protein as a protective antigen and vaccine candidate is warranted.

Adenylyl Cyclase Inhibitors↗
PubMed1987

Further characterization of the subunits of the receptor with high affinity for immunoglobulin E.

The alpha, beta, and gamma subunits of the receptor with high affinity for immunoglobulin E were isolated and their compositions assessed by direct amino acid analysis and by incorporation of radioactive precursors. The compositions show no unusual features other than a rather high content of tryptophan in the alpha chain as assessed from the incorporation studies. The results combined with future sequence data will permit unambiguous determination of the multiplicity of the chains in the receptor. Chymotryptic peptide maps of the extrinsically iodinated subunits show several similar peptides, particularly for alpha and beta. However, these putative homologies were not apparent when tryptic maps of the biosynthetically ([3H]leucine) labeled subunits were analyzed.

Amino Acids↗
PubMed1987

Biosynthesis of human C-reactive protein in cultured hepatoma cells is induced by a monocyte factor(s) other than interleukin-1.

An in vitro cell culture system utilizing continuous human liver cells has been developed which, upon specific induction, will respond by synthesizing, de novo, the prototype acute phase reactant, C-reactive protein (CRP). Induction of CRP in vitro is not brought about by the types of hormones, steroids, and chemicals which affect other acute phase proteins. In particular, interleukin-1 thought to be directly responsible for acute phase induction is not found to be active. Direct testing of other purified biological response modifiers, i.e. alpha, beta, and gamma-interferon, interleukin-2, and tumor necrosis factor, demonstrates no inducing activity. However, we find that human peripheral blood monocytes, stimulated by endotoxin, produce a factor(s) which directly induces CRP synthesis in hepatoma cells. In addition, the human promyelocyte-like cell line HL-60 in the presence of phorbol ester and certain T-cell lines containing human retroviruses also produce this CRP-inducing factor(s). Isolation and partial purification of the CRP-inducing factor(s) indicate that it is a protein(s) with a molecular weight of approximately 30,000.

C-Reactive Protein↗
PubMed1987

Preparation of phosphorylcholine derivatives of bovine serum albumin and their application to the affinity chromatography of C-reactive protein.

A simple method for the preparation of phosphorylcholine derivatives of bovine serum albumin (PC-BSA) by reductive alkylation of the amino groups of bovine serum albumin with choline phosphoryl glycoaldehyde is described. Choline phosphoryl glycoaldehyde was generated by periodate oxidation of glyceryl phosphorylcholine. PC-BSA was immobilized on SH-derivatized Toyopearl HW 65 by reacting the single SH group of PC-BSA with a bismaleimido reagent and then coupling maleimidated PC-BSA to the thiolated gel. The affinity purification of C-reactive protein (CRP), which is based on the Ca2+-dependent affinity of CRP for the phosphorylcholine residue of PC-BSA, was readily accomplished using the PC-BSA Toyopearl HW 65 column. The resulting CRP preparation from serum and pleural fluid was homogeneous as assessed by native polyacrylamide gel electrophoresis. PC-BSA derivatives were also shown to be reactive with Limulus polyphemus CRP.

C-Reactive Protein↗
PubMed1987

Purification and characterization of serotype 6 fimbriae from Bordetella pertussis and comparison of their properties with serotype 2 fimbriae.

Fimbriae were removed from Bordetella pertussis (serotype 1.3.6) by mechanical shearing and purified by precipitation with ammonium sulfate, pH-dependent precipitation at pH 7.4, followed by two successive extractions of the precipitated fimbriae with 4 M urea. By electron microscopy, the precipitated fimbriae appeared as aggregated bundles of long, relatively straight filaments which were disaggregated to individual flexuous filaments at pH 10.5. These purified fimbriae were identified as serotype 6 agglutinogens, since antibody to the purified fimbriae agglutinated B. pertussis strains serotyped as 1.3.6, 1.2.3.6, or 1.2.3.4.6 but did not agglutinate strains of serotype 1.2.3.4, 1.2.3, or 1.3. In contrast, antibody to serotype 2 fimbriae only agglutinated B. pertussis strains containing serotype 2 agglutinogen. Purified type 6 and 2 fimbriae were found to be weakly cross-reactive by enzyme-linked immunosorbent assay, using polyclonal antibody to each type of fimbria. In an immunoblot assay, polyclonal antibodies to a 22,000-dalton subunit of fimbriae from B. bronchiseptica reacted strongly with the type 2 fimbrial subunit of B. pertussis, but only weakly with the type 6 subunit. When subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein subunit of the type 6 fimbriae migrated with a molecular weight of 21,500, whereas the type 2 fimbrial subunit had a molecular weight of 22,000. The two types of subunits had similar amino acid compositions and showed amino-terminal sequence homology in 15 of 21 amino acids. The amino-terminal amino acid sequences of the B. pertussis fimbriae were distinct from those reported for fimbriae from other gram-negative bacteria. Neither the type 6 nor the type 2 fimbriae caused hemagglutination when assayed with several types of erythrocytes.

Amino Acid Sequence↗
PubMed1986

Rubella virus cDNA. Sequence and expression of E1 envelope protein.

A cDNA clone encoding the entire E1 envelope protein (410 amino acid residues) and a portion of the C-terminal end of the E2 envelope protein of the rubella virus has been isolated and characterized. DNA sequence analysis has revealed a region 20 nucleotides in length at the 3' end of the cloned cDNA which may be a replicase recognition site or a recognition site for encapsidation. The proteolytic cleavage site between the E1 and E2 proteins was localized based on the known amino-terminal sequence of the isolated E1 protein (Kalkkinen, N., Oker-Blom, C., and Pettersson, R. F. (1984) J. Gen. Virol. 65, 1549-1557) and the deduced amino acid sequence. The mature E1 protein is preceded by a set of 20 highly hydrophobic amino acid residues possessing characteristics of a signal peptide. This "signal peptide" is flanked on both sides by typical protease cleavage sites for trypsin-like enzyme and signal peptidase. The presence of a leader sequence in the E1 protein precursor may facilitate its translocation through the host cell membrane. The E1 protein of rubella virus shows no significant homology with alphavirus E1 envelope proteins. However, a stretch of 39 amino acids in the E1 protein of rubella virus (residues 262-300) was found to share a significant homology with the first 39 residues of bovine sperm histone. The position of 4 half-cystines and 8 arginines overlaps. The E1 protein of rubella virus has been successfully expressed in COS cells after transfecting them with rubella virus cDNA in simian virus 40-derived expression vector. This protein is antigenically similar to the one expressed by cells infected with rubella virus.

Amino Acid Sequence↗