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Biomedical subjects

J T Yang

Publications and source records attributed to J T Yang.

At least 55 records · Page 3Linked to original sources
PubMed1993

Eosinophilic granuloma of the skull--report of four cases.

Eosinophilic granuloma is a benign form of the three clinical variants of histiocytosis-X. It is an uncommon, localized granulomatous process, forming solitary or multiple lytic skeletal lesions, and primarily involving male children and young adults. We studied 4 cases, three boys and one girl, aged 4, 8, 27 and 12 years, in Chang Gung Memorial Hospital from 1981 to 1990. Their common presentation was the rapidly growing, painful, and protruding scalp masses without neurological deficits. En bloc excisions of these lesions were performed. Then radiotherapy was given postoperatively in two cases, chemotherapy in one, and no adjuvant treatment in one. During their follow-up period of 2 to 10 years, all 4 patients were well, without evidence of recurrence.

Adult↗
PubMed1992

Beta-II conformation of all-beta proteins can be distinguished from unordered form by circular dichroism.

The CD spectrum of certain all-beta globular proteins resembles that of unfolded proteins with a characteristic negative band around 200 nm. The conformation of this class is tentatively termed beta-II, which had two features that were absent for unfolded proteins. First, beta-II proteins usually had CD bands due to aromatic side groups in the near-ultraviolet region. Second, the CD intensities both in the far- and in the near-uv region of these compact and rigid proteins usually showed a sharp transition upon thermal denaturation, whereas those of an unordered form changed linearly with rising temperature.

Chymotrypsin↗
PubMed1992

Conformational stability of porcine serum transferrin.

The conformation of porcine serum ferric transferrin (Tf) and its stability against denaturation were studied by circular dichroism. Tf was estimated to have 19-24% alpha-helix and 50-55% beta-sheet based on the methods of Chang et al. (Chang, C.T., Wu, C.-S.C., & Yang, J.T., 1978, Anal. Biochem. 91, 13-31) and Provencher and Glöckner (Provencher, S.W. & Glöckner, J., 1981, Biochemistry 20, 33-37). Removal of the bound ferric ions (apo-Tf) did not alter the overall conformation, but there were subtle changes in local conformation based on its near-UV CD spectrum. The Tfs were stable between pH 3.5 and 11. Denaturation by guanidine hydrochloride (Gu-HCl) showed two transitions at 1.6 and 3.4 M denaturant. The process of denaturation by acid and base was reversible, whereas that by Gu-HCl was partially reversible. The irreversible thermal unfolding of Tfs began at temperatures above 60 degrees C and was not complete even at 80 degrees C. The bound irons (based on absorbance at 460 nm) were completely released at pH < 4 or in Gu-HCl solution above 1.7 M, when the protein began to unfold, but they remained intact in neutral solution even at 85 degrees C. The NH2- and COOH-terminal halves of the Tf molecule obtained by limited trypsin digestion had CD spectra similar to the spectrum of native Tf, and the COOH-terminal fragment had more stable secondary structure than the NH2-terminal fragment.

Animals↗
PubMed1992

Conformation of concanavalin A and its fragments in aqueous solution and organic solvent-water mixtures.

The conformations of concanavalin A (con A), an all-beta protein, and its three CNBr-cleaved fragments were studied by CD. Con A in buffer showed a 197 nm maximum and a 223 nm minimum, which were red-shifted by 6-7 nm from those of regular all-beta proteins and beta-sheet of (Lys)n. Fragment 1 (residue 1-42) resembled an unordered form with a CD maximum at 200 nm, but fragments 2 (residues 43-129) and 3 (residues 130-237) showed a regular CD spectrum with two extrema at 192-193 nm (+) and 214-216 nm (-). Equimolar mixture of the three fragments showed some degree of interaction, but did not reconstitute the conformation of native con A, probably because of the loss of bound Ca2+ and Mn2+ ions in the fragments. In ethanol-, methanol-, and dioxane-water mixed solvents, con A and its fragments remained as beta-sheet. In contrast, addition of trifluoroethanol and sodium dodecyl sulfate induced alpha-helix at the expense of beta-sheet for con A and its fragments in aqueous solution. In 80% trifluoroethanol, the induced helicities exceeded their sequence-predicted helix-potentials, but in 10 mM sodium dodecyl sulfate the helicities agreed well with corresponding predictions.

Buffers↗
PubMed1991

Some problems of CD analyses of protein conformation.

The circular dichroic (CD) spectra of 16 reference proteins were analyzed by the Provencher-Glöckner method (Biochemistry 20, 31, 1981) with the lower-wavelength limit raised from 190 to 235 nm at 5-nm intervals. Fifty-one data points at 1-nm intervals were taken between 190 and 240 nm. Variations of the correlation coefficients (r) and root-mean-square (RMS) deviations between X-ray diffraction results and CD analyses showed no definite trend with shorter wavelength ranges. The CD spectra (190-240 nm) were also analyzed by assigning the secondary structure of X-ray results according to the Levitt-Greer method (J. Mol. Biol. 114, 181, 1977) and the Kabsch-Sander method (Biopolymers 22, 2577, 1983). The r and RMS values based on the Levitt-Greer assignment were good and comparable to those based on the secondary structure given by crystallographers, but the Kabsch-Sander assignment seemed to give unsatisfactory results. The choice of reference proteins remains one of the uncertainties in the CD analysis. The five most significant orthogonal spectra (190-240 nm) calculated from the 16 reference proteins and those based on another 16 proteins used by Hennessey and Johnson (Biochemistry 20, 1085, 1981) were similar to each other, but different in intensities. These methods still cannot recognize a failed analysis of unknown proteins without X-ray results to check their reliability.

Circular Dichroism↗
PubMed1991

Basic fibroblast growth factor is a beta-rich protein.

The conformation of the 153-residue form of human basic fibroblast growth factor (bFGF) was studied with circular dichroism (CD) and sequence prediction methods. The far-UV CD spectrum with a minimum at 202 nm resembled that of an unordered polypeptide/protein or a protein rich in distorted antiparallel beta-sheets. Analysis of the CD spectrum by the least-squares method of Chang et al. (1978) and the CONTIN program of Provencher and Glöckner (1981) suggested that about one half of the molecule consisted of beta-sheet and there was no alpha-helix. These estimates agreed with the prediction by the sequence method of Garnier et al. (1978) using decision constants based on CD results. bFGF had an unusual CD band at 187 nm, which disappeared upon ionization of Tyr side chains at pH 11.7. It also had another unusual property of irreversibly converting the CD spectrum to a helix-like one with a double minimum at 205 and 215 and a maximum at 189 nm upon heating the solution to above 55 degrees C. The helicity was also enhanced in trifluoroethanol and in sodium dodecyl sulfate. The mutant bFGF in which cysteines 76 and 94 were replaced by serine residues had essentially the same properties as the wild-type.

Amino Acid Sequence↗
PubMed1991

Quantitative analysis of nonoxynol-9 in blood.

At present, there is little data on the level of nonoxynol-9 (N-9) in blood. Using routine methods, the rate of recovery for N-9 in blood has been confirmed to be very low. A method determining N-9 levels in urine has been found but obviously a more efficient method for determining N-9 levels in blood and urine is still needed. Here, a method of determining N-9, a nonionic surfactant spermicide, in blood is presented. The analytical procedure is a simple and convenient method. Using HPLC, UV detector, ODS reversed-phase column, methanol-water mobile phase, NaCl salting-out and two extractions with benzene, a rate of recovery for N-9 of 93 +/- 4% has been obtained.

Administration, Intravaginal↗
PubMed1990

Evidence that the head of kinesin is sufficient for force generation and motility in vitro.

Kinesin is a mechanochemical protein that converts the chemical energy in adenosine triphosphate into mechanical force for movement of cellular components along microtubules. The regions of the kinesin molecule responsible for generating movement were determined by studying the heavy chain of Drosophila kinesin, and its truncated forms, expressed in Escherichia coli. The results demonstrate that (i) kinesin heavy chain alone, without the light chains and other eukaryotic factors, is able to induce microtubule movement in vitro, and (ii) a fragment likely to contain only the kinesin head is also capable of inducing microtubule motility. Thus, the amino-terminal 450 amino acids of kinesin contain all the basic elements needed to convert chemical energy into mechanical force.

Adenosine Triphosphatases↗
PubMed1990

Thermal unfolding of helices of a C-peptide analogue of ribonuclease A in sodium dodecyl sulfate solution.

The conformation of a 13-residue C-peptide analogue of ribonuclease A, suc-AET-AAAKFLRAHA-CONH2, in NaDodSO4 solutions with respect to temperature was studied with CD. The equilibrium constant of unfolding yielded a straight line in a van't Hoff plot. In 10 mM NaDodSO4, delta G mu = 120 cal/mol, delta H mu = 700 cal/mol, and delta S mu = 2.0 entropy units all on per helical residue. These values compared fairly well with the thermodynamic parameters of the uncharged helix-coil transition of (Glu)n in 0.1 M NaCl based on the theories of Zimm and Bragg and Zimm and Rice. The peptide was not unfolded at 75 degrees C completely. Even in water without surfactant it was not a "random coil."

Amino Acid Sequence↗
PubMed1990

Opioid-binding protein (OBCAM) is rich in beta-sheets.

Based on circular dichroism (CD) and the sequence-predictive method, the opioid-binding cell adhesion molecule (OBCAM) consisted of one half beta-sheets and one fourth alpha-helices. This is consistent with significant sequence homology of the protein to several members of the immunoglobulin (Ig) superfamily, particularly cell adhesion molecules, which are rich in beta-sheets. Hydropathy analysis suggests that hydrophobic and hydrophilic regions were evenly distributed along the sequence, but the NH2- and COOH-termini were hydrophobic. Hydrophobic moments and Fourier-transform amphipathic analyses further suggest that residues 23-30 and 83-93 were amphipathic beta-sheets. The overall conformation of OBCAM was unaltered by adding linoleic acid, which is required for opioid ligand binding.

Animals↗
PubMed1990

Conformation of acetylcholine receptor in the presence of agonists and antagonists.

The conformations of acetylcholine receptor from Torpedo californica in the absence and presence of agonists, antagonists, and local anesthetics were studied by circular dichroism (CD). Without ligands, the receptor had about 40% helix, 20% beta-sheets, and 10% beta-turns as analyzed from its far-UV CD spectrum. Its near-UV CD spectrum resembled that of acetylcholinesterase from the same source. None of the ligands studied altered the far-UV spectrum of the receptor. However, in the near-UV region, carbamylcholine and acetylcholine shifted the Phe and Tyr bands of AChR to less negative, whereas hexamethonium changed the Tyr bands to more negative, indicating that the site of binding of agonists and antagonists and their effect on the conformation of the receptor may be different. Decamethonium, procaine, and lidocaine had no effect on both the far- and near-UV CD spectra of acetylcholine receptor.

Anesthetics, Local↗
PubMed1989

Identification of globular mechanochemical heads of kinesin.

Kinesin is a mechanoenzyme which uses energy liberated from ATP hydrolysis to transport particles towards the 'plus ends' of microtubules. The enzyme consists of two polypeptide heavy chains of relative molecular mass (Mr) approximately 110,000-140,000 (110K-140K) plus copurifying light chains; these polypeptides are arranged in a structure consisting of two globular heads attached to a fibrous stalk which terminates in a 'feathered' tail. Here we report that a function-disrupting monoclonal antikinesin, which binds to the 45K fragment of the kinesin heavy chain, recognizes an epitope located towards the N-terminal end of the heavy chain, and decorates the two globular heads lying at one end of the intact molecules (one antibody per head). The results show that the two heavy chains of native kinesin are arranged in parallel, and that the 45K fragments, which display nucleotide-sensitive interactions with microtubules, represent mechanochemical 'heads' located at the N-terminal regions of the heavy chains. Thus, it is likely that the kinesin heads are analogous to the subfragment-1 domains of myosin.

Animals↗
PubMed1989

A three-domain structure of kinesin heavy chain revealed by DNA sequence and microtubule binding analyses.

The structure and function of kinesin heavy chain from D. melanogaster have been studied using DNA sequence analysis and analysis of the properties of truncated kinesin heavy chain synthesized in vitro. Analysis of the sequence suggests the existence of a 50 kd globular amino-terminal domain that contains an ATP binding consensus sequence, followed by another 50-60 kd domain that has sequence characteristics consistent with the ability to fold into an alpha helical coiled coil. The properties of amino- and carboxy-terminally truncated kinesin heavy chains synthesized in vitro reveal that a 60 kd amino-terminal fragment has the nucleotide-dependent microtubule binding activities of the intact kinesin heavy chain, and hence is likely to be a "motor" domain. Finally, the sequence data indicate the presence of a small carboxy-terminal domain. Because it is located at the end of the molecule away from the putative "motor" domain, we propose that this domain is responsible for interactions with other proteins, vesicles, or organelles. These data suggest that kinesin has an organization very similar to that of myosin even though there are no obvious sequence similarities between the two molecules.

Adenosine Triphosphatases↗
PubMed1989

Preparation of clear solutions of reconstituted acetylcholinesterase. Effect of ionic strength and lipid/protein ratio.

The detergent-soluble globular dimer of acetylcholinesterase from Torpedo californica was reconstituted through dialysis into preformed egg phosphatidylcholine vesicles. The formation of the enzyme-lipid complexes depended on the ionic strength of the dialysis buffer as well as the molar lipid/protein ratio (R). The enzyme was unstable at I less than 0.05; increasing the ionic strength increased the size of the complex. A too low R value (e.g. 1000) would promote self-aggregation of the enzyme and produce heterogeneous complexes, especially at high I values. On the other hand, a too high R value (e.g. greater than 5000) favored the formation of large enzyme-lipid complexes; their solutions were too turbid for optical studies. The enzyme reconstituted at I = 0.07 and R = 4000 gave a clear solution and showed no artifacts due to light scattering. The conformation based on circular dichroism and enzymatic activity of the detergent-soluble enzyme were unchanged upon reconstitution. The reconstituted enzyme in lipid vesicles seemed to be slightly more stable against thermal denaturation than the protein in sodium cholate solution.

Acetylcholinesterase↗
PubMed1989

Transfer of foreign genes by electroporation and their expression in mammalian cells.

Expression of foreign genes transferred into mammalian cells by electroporation has been studied. The pX1TK gene, pSV2Neo gene and pUCEJ oncogene have been introduced into MLTK- cells and NIH/3T3 cells, respectively. Stable transformation transient expression of TK gene by MLTK- cells as well as stable and malignant transformation of NIH/3T3 cells have been obtained. Transient expression frequency is about 80% and stable transformation frequency is about 10(-4). Integration of foreign genes into the cellular genome was verified with molecular hybridization. Tumor development was observed after inoculation of transformed cells into nude mice.

Animals↗
PubMed1989

Tacrine protection of acetylcholinesterase from inactivation by diisopropylfluorophosphate: a circular dichroism study.

Tacrine (1,2,3,4-tetrahydro-9-aminoacridine) showed an apparent noncompetitive inhibition of Torpedo acetylcholinesterase (AChE) with a dissociation constant, Ki, of 8.5 nM. It altered the CD bands of AChE in the near-UV region, which monitor the local conformation of aromatic side groups, but not those in the far-UV region, which measure the secondary structure. An extrinsic CD band was induced at 348 nm, with a molar ellipticity of 35,000 deg cm2 dmol-1 (bases on tacrine), when each AChE subunit (Mr = 67,000) was saturated with one tacrine (mol/mol). With this band as a probe, the bound tacrine could be displaced by edrophonium or decamethonium, both of which are known to bind to the anionic site at the active center of AChE, but not by propidium, which binds to the peripheral site of the enzyme. Tacrine protected AChE from inactivation by diisopropylfluorophosphate (DFP). AChE completely lost its enzymatic activity when 1 mol of DFP was bound per mol of subunit upon incubation of 7 microM AChE (subunit) with 100 microM DFP for 40 min, but tacrine-treated AChE retained 60% of its activity and bound only 0.2 mol of DFP per mol of subunit under similar conditions. The corresponding CD, at 348 nm, of the AChE-tacrine-DFP complex increased or decreased gradually, depending on the order of addition of tacrine and DFP, and reached an equilibrium value (80% of its original) after 2 days. The difference absorption spectrum of the AChE-tacrine-DFP complex was the same as that of the AChE-tacrine complex. These results suggest that the protective effect of tacrine may be due to steric hindrance at the esteratic site of the enzyme.

Acetylcholinesterase↗