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J W Cheng

Publications and source records attributed to J W Cheng.

54 records · Page 3Linked to original sources
PubMed1995

Spindle dynamics and cell cycle regulation of dynein in the budding yeast, Saccharomyces cerevisiae.

We have used time-lapse digital- and video-enhanced differential interference contrast (DE-DIC, VE-DIC) microscopy to study the role of dynein in spindle and nuclear dynamics in the yeast Saccharomyces cerevisiae. The real-time analysis reveals six stages in the spindle cycle. Anaphase B onset appears marked by a rapid phase of spindle elongation, simultaneous with nuclear migration into the daughter cell. The onset and kinetics of rapid spindle elongation are identical in wild type and dynein mutants. In the absence of dynein the nucleus does not migrate as close to the neck as in wild-type cells and initial spindle elongation is confined primarily to the mother cell. Rapid oscillations of the elongating spindle between the mother and bud are observed in wild-type cells, followed by a slower growth phase until the spindle reaches its maximal length. This stage is protracted in the dynein mutants and devoid of oscillatory motion. Thus dynein is required for rapid penetration of the nucleus into the bud and anaphase B spindle dynamics. Genetic analysis reveals that in the absence of a functional central spindle (ndcl), dynein is essential for chromosome movement into the bud. Immunofluorescent localization of dynein-beta-galactosidase fusion proteins reveals that dynein is associated with spindle pole bodies and the cell cortex: with spindle pole body localization dependent on intact microtubules. A kinetic analysis of nuclear movement also revealed that cytokinesis is delayed until nuclear translocation is completed, indicative of a surveillance pathway monitoring nuclear transit into the bud.

Anaphase↗
PubMed1994

Solution structure of FK506 bound to the R42K, H87V double mutant of FKBP-12.

The binding of the FK506/FKBP-12 complex to calcineurin (CN), its putative target for immunosuppression, involves recognition of solvent-exposed regions of the ligand as well as FKBP-12 residues near the active site. The R42K, H87V double mutation of FKBP-12 decreases the CN affinity of the complex by 550-fold [Aldape, R. A., Futer, O., DeCenzo, M. T., Jarrett, B. P., Murcko, M. A., & Livingston, D. J. (1992) J. Biol. Chem. 267, 16029-16032]. This work reports the solution structure of 13C-labeled FK506 bound to R42K, H87V FKBP-12. Assignments and NOE measurements at three mixing times were made from inverse-detected 1H-13C NMR experiments. Structures were calculated by several different methods, including distance geometry, restrained molecular dynamics, and molecular dynamics with time-averaged restraints. The NMR structures of the ligand are very well defined by the NOE restraints and differ slightly from the X-ray structure in regions that are involved in crystal packing. Comparison with the NMR structure of FK506 bound to wild-type FKBP-12 reveals that the R42K, H87V mutation causes the ligand backbone near C16 to move by 2.5 to 4.5 A, reorients 15-MeO by 90 degrees, and shifts 13-MeO by approximately 1.5 A. FK506 appears to undergo a concerted, mutationally induced shift in the binding pocket, with the greatest changes occurring in the effector region of the drug. The altered effector conformation of mutant-bound FK506 may perturb interactions between the drug and CN, thus accounting for the effect of the double mutation upon the CN inhibitory activity of the complex.

Calcineurin↗
PubMed1994

DNA sequence GCGAATGAGC containing the human centromere core sequence GAAT forms a self-complementary duplex with sheared G.A pairs in solution.

The DNA sequence dGCGAATGAGC has a well-resolved, two-dimensional nuclear Overhauser (NOESY) spectrum that is suitable for high quality solution structure determination by NMR methods; in solution this sequence forms a stable self-complementary duplex containing sheared G.A base-pairs. A total of 220 distance constraints derived from time-dependent NOE measurements were collected and refined by repeated back-calculation of the NOESY spectra. Distance information from imino proton studies and from exclusive two-dimensional correlated spectroscopy (E. COSY) and/or linewidth analysis was included in the structure calculation using the program DSPACE 4.2, followed by restrained energy minimization with the program DISCOVER using the AMBER force field. The energies of the distance geometry (DG) structures decreased rapidly in the first few cycles and approached -510 +/- 3 kcal after 1000 cycles of conjugate gradient minimization (about 540 kcal lower than in the initial DG structures). All 15 final DG structures converged to a single family of closely related structures with pair-wise r.m.s.d. values of 0.96 +/- 0.34 A, which was further reduced by energy minimization to 0.70 +/- 0.35 A. Rather unusual structural features of the duplex are revealed in the final structures. The results indicate that, in addition to normal sequences with standard base-pairing, unusual nucleic acid structures can also be determined in solution with quite high precision by NMR/distance geometry methods.

Base Composition↗
PubMed1994

15N NMR relaxation studies of the FK506 binding protein: dynamic effects of ligand binding and implications for calcineurin recognition.

Backbone dynamics of the ligand- (FK506-) bound protein FKBP-12 (107 amino acids) have been examined using 15N relaxation data derived from inverse-detected two-dimensional 1H-15N NMR spectra. A model free formalism [Lipari & Szabo (1982) J. Am. Chem. Soc. 104, 4546-4559] was used to derive the generalized order parameter (S2), the effective correlation time for internal motions (tau e), and the chemical-exchange line width (R(ex)) based on the measured 15N relaxation rate constants (R1, R2) and 1H-15N heteronuclear NOEs. The final optimized overall correlation time (tau m) was 9.0 ns. The average order parameter (S2) describing the amplitude of motions on the picosecond time scale was found to be 0.88 +/- 0.04, indicating that internal flexibility is restricted along the entire polypeptide chain. In contrast to results obtained for uncomplexed FKBP, the 80's loop (residues 82-87) surrounding the ligand binding site was found to be rigidly fixed, indicating that internal motions at this site are damped significantly due to stabilizing noncovalent interactions with the FK506 molecule. Structural implications of these differences in picosecond mobility as well as possible implications for calcineurin recognition are discussed.

Binding Sites↗
PubMed1994

Should all patients with dilated cardiomyopathy receive chronic anticoagulation?

OBJECTIVE: To review the mechanism of intracardiac thrombus formation, describe the risk of systemic thromboembolization, critically evaluate the literature regarding the use of oral anticoagulation, review the American College of Chest Physicians (ACCP) guidelines, and provide recommendations for oral anticoagulation in patients with dilated cardiomyopathy (DCM). DATA SOURCES: English language clinical studies, abstracts, and review articles pertaining to oral anticoagulation and DCM. STUDY SELECTION AND DATA EXTRACTION: Relevant human studies examining the role of anticoagulation for preventing systemic thromboembolism in patients with DCM. DATA SYNTHESIS: Potential mechanisms and risk factors for systemic thromboembolization in patients with DCM are discussed. Studies evaluating the benefits and risks of chronic oral anticoagulation to prevent systemic thromboembolization are critiqued. Recommendations for oral anticoagulation in patients with DCM from the second and third ACCP Consensus Conferences are reviewed. Suggestions for study design of a new clinical trial are presented. CONCLUSIONS: The second ACCP Consensus Conference recommended chronic anticoagulation for all patients with DCM, but the third conference did not address this issue. Review of demographic data from clinical trials demonstrates reluctance to use anticoagulation in all patients with DCM because of lack of support from prospective, controlled trials. Recent data from large clinical trials suggest that the risk of systemic embolization may be lower than previously believed. We recommend the use of chronic anticoagulation with warfarin in patients for whom the risk of embolism is greater than the risk of major bleeding, such as those with atrial fibrillation or previous systemic embolization. Lack of compliance with ACCP guidelines suggests that clinicians require additional information regarding the bleeding risk and systemic embolization rate reduction of chronic anticoagulation in patients with DCM who remain in normal sinus rhythm. It is only through prospective, controlled trials that this risk/benefit ratio to prevent systemic embolism in patients with DCM can be established.

Administration, Oral↗
PubMed1993

15N NMR relaxation studies of the FK506 binding protein: backbone dynamics of the uncomplexed receptor.

Backbone dynamics of the major tacrolimus (FK506) binding protein (FKBP-12, 107 amino acids) have been studied using 15N relaxation data derived from proton-detected two-dimensional 1H-15N NMR spectroscopy. 15N spin-lattice relaxation rate constants (R1), spin-spin relaxation rate constants (R2), and heteronuclear NOEs were determined for over 85% of the backbone amide 15N nuclei. A model free formalism [Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4546-4559; Lipari, G., & Szabo, A. (1982) J. Am. Chem. Soc. 104, 4559-4570] was used to derive values for the generalized order parameter (S2), the effective correlation time for internal motions (tau e), and the chemical exchange line width (Rex) for each N-H bond vector. The final optimized overall correlation time (tau m) was 9.2 ns. The average order parameter (S2) describing the amplitude of motions on the picosecond time scale was found to be 0.88 +/- 0.06. Motions on the picosecond time scale are restricted at the N and C termini, consistent with previous NMR structural studies indicating well-defined beta-strands in these regions. With the exception of the flap region from resides 82 to 87, no regions appear to be significantly disordered on the picosecond time scale. Residues in several regions of the protein exhibit high Rex terms, indicating possible motions on the millisecond to microsecond time scale due to chemical exchange and/or conformational averaging effects. Possible effects of tacrolimus (FK506) binding on FKBP-12 dynamics are discussed in the context of previously determined solution structures for FKBP-12 in the uncomplexed [Michnick et al. (1991) Science 252, 836-839; Moore et al. (1991) Nature 351, 248-250] and complexed [Meadows et al. (1993) Biochemistry 32, 754-765] states.

Amides↗
PubMed1992

Base pairing geometry in GA mismatches depends entirely on the neighboring sequence.

We have synthesized nine self-complementary DNA oligomers containing different flanking sequences adjacent to a pair of contiguous GA mismatches, and have used high resolution nuclear magnetic resonance (n.m.r.) to investigate the GpA phosphodiester backbone conformation and mismatch pairing schemes in these duplexes. We found dramatic effects of the flanking base pair on the hydrogen bonding and backbone conformation, which appear to be coupled. Thus the Ganti-Aanti base pairing scheme in a NAGATN sequence switches to a more stable sheared GA base pairing scheme in a NCGAGN or NTGAAN context, while no duplex is formed (or only GA bulges occur) when NAGATN is changed to NGGACN. Furthermore, the more stable sheared GA pairing in NPyGAPuN sequences is associated with a BII rather than BI backbone conformation for the phosphodiester between the adjacent mismatched GA pairs. The overall stability of these adjacent GA mismatches as measured by imino proton n.m.r. studies is Py-GA-Pu > A-GA-T > G-GA-C.

Base Composition↗
PubMed1992

Solution structure of [d(GCGTATACGC)]2.

The solution structure of the alternating pyrimidine-purine DNA duplex [d(GCGTATACGC)]2 has been determined using two-dimensional nuclear magnetic resonance techniques and distance geometry methods. Backbone distance constraints derived from experimental nuclear Overhauser enhancement and J-coupling torsion angle constraints were required to adequately define the conformation of the inter-residue backbone linkages and to avoid underwinding of the duplex. The distance geometry structures were further refined by back-calculation of the two-dimensional nuclear Overhauser enhancement spectra to correct spin-diffusion distance errors. Fifteen final structures for [d(GCGTATACGC)]2 were generated from the refined experimental distance bounds. These structures all exhibit fully wound B-form geometry with small penalty values (< 1.5 A) against the distance bounds and small pair-wise root-mean-square deviation values (typically 0.6 A to 1.5 A). The final structures exhibit positive base-pair inclination with respect to the helix axis, a marked alternation in rise and twist, and are shorter and wider than classical fiber B-form DNA. The purines were found to adopt a sugar pucker close to the C-2'-endo conformation while pyrimidine sugars exhibited significantly lower pseudorotation phase angles in the C-1'-exo to C-2'-endo range. The minor groove cross-strand steric clashes at pyrimidine-purine steps that would exist in pure B-DNA are attenuated by an increased rise at these steps (and an increased roll angle at TpA steps). Concomitantly the backbone torsion angles of the pyrimidine moieties have larger gamma values, larger epsilon values, and smaller zeta values than the purines. The structures generated by distance geometry methods were also compared with those obtained from restrained molecular dynamics with empirical force-field potentials. The results indicate that the nuclear magnetic resonance/distance geometry approach alone is capable of elucidating most of the salient structural features of double-stranded helical nucleic acids in solution without resorting to empirical energy potentials and without using any structural assumptions from crystallographic data.

Base Sequence↗
PubMed1992

Solution structure of [d(ATGAGCGAATA)]2. Adjacent G:A mismatches stabilized by cross-strand base-stacking and BII phosphate groups.

The solution structure of a rather unusual B-form duplex [d(ATGAGCGAATA)]2 has been determined using two-dimensional nuclear magnetic resonance (2D-NMR) and distance geometry methods. This sequence forms a stable ten base-pair B-form duplex with 3' overhangs and two pairs of adjacent G:A mismatches paired via a sheared hydrogen-bonding scheme. All non-exchangeable protons, including the stereo-specific H-5'S/H-5'R of the 3G and 7G residues, were assigned by 2D-NMR. The phosphorus spectrum was assigned using heteronuclear correlation with H-3' and H-4' reasonances. The complete assignments reveal several unusual nuclear Overhauser enhancements (NOEs) and unusual chemical shifts for the neighboring G:A mismatch pairs and their adjacent nucleotides. Inter-proton distances were derived from time-dependent NOEs and used to generate initial structures, which were further refined by iterative back-calculation of the two-dimensional nuclear Overhauser enhancement spectra; 22 final structures were calculated from the refined distance bounds. All these final structures exhibit fully wound helical structures with small penalty values against the refined distance bounds and small pair-wise root-mean-square deviation values (typically 0.5 A to 0.9 A). The two helical strands exchange base stacking at both of the two G:A mismatch sites, resulting in base stacking down each side rather than down each strand of the twisted duplex. Very large twist angles (77 degrees) were found at the G:A mismatch steps. All the final structures were found to have BII phosphate conformations at the adjacent G:A mismatch sites, consistent with observed downfield 31P chemical shifts and Monte-Carlo conformational search results. Our results support the hypothesis that 31P chemical shifts are related to backbone torsion angles. These BII phosphate conformations in the adjacent G:A mismatch step suggest that hydrogen bonding of the G:A pair G-NH2 to a nearby phosphate oxygen atom is unlikely. The unusual structure of the duplex may be stabilized by strong interstrand base stacking as well as intrastrand stacking, as indicated by excellent base overlap within the mismatch stacks.

Base Composition↗
PubMedDate not supplied

Calcium channel blockers: association with myocardial infarction, mortality, and cancer.

The purpose of this study was to review the results of trials assessing the association between the use of calcium channel blockers (CCBs) and mortality, myocardial infarction (MI), and cancer. Possible mechanisms of such relationships are discussed and recommendations regarding the use of CCBs made. Since 1995, 10 controversial studies have been published that associate the use of CCBs with an increased risk of mortality, MI, and cancer; these findings have caused widespread anxiety and frustration among patients and physicians. For health care professionals to properly advise patients, the facts surrounding this controversy should be reviewed. To do this, we reviewed and assessed English-language clinical studies, abstracts, editorials, and review articles pertaining to the use of CCBs and mortality, MI, and cancer in humans. The designs of ongoing prospective, randomized studies are discussed. Based on current published studies, the US Food and Drug Administration has agreed to a label warning against off-label use of short-acting nifedipine in patients with hypertension, acute MI, or nonvasospastic unstable angina. Practitioners should exercise caution when prescribing CCBs, especially to high-risk patients (e.g., those with congestive heart failure or clinical or subclinical coronary artery disease). When possible, long-acting CCBs should be used.

Animals↗
PubMedDate not supplied

Solution conformation of a peptide corresponding to residues 151-172 of HIV-1 integrase using NMR and CD spectroscopy.

The solution structure of a synthetic peptide corresponding to residues 151-172 of HIV-1 integrase has been determined by NMR and CD spectroscopy. Residues 151-172 of HIV-1 integrase were predicted to be an alpha-helix and to be responsible for the oligomerization of HIV-1 integrase. Two-dimensional 1H NMR and CD studies indicate that this synthetic peptide adopts an amphipathic alpha-helical conformation in TFE-containing solution. However, concentration-dependent CD studies reveal that this peptide motif does not form dimers or oligomers in solution as predicted. These results are in agreement with the crystal structure of the catalytic domain of HIV-1 integrase reported recently.

Amino Acid Sequence↗
PubMedDate not supplied

Patient preferences for thrombolytic therapy in acute myocardial infarction.

BACKGROUND: Despite extensive professional debate regarding the optimal thrombolytic therapy strategy in acute myocardial infarction (AMI), patient preferences have not been explored. METHODS: Preferences among patients with known or suspected coronary artery disease for treatment with tissue plasminogen activator (tPA) or streptokinase (SK) for AMI were determined using a questionnaire presenting GUSTO-1 trial and drug cost data. Preferences were based on consideration of 30-day mortality (M) alone, hemorrhagic stroke rate (SR) alone, overall preference (M + SR), drug acquisition costs, and the estimated annual costs of using a single agent to treat all AMIs. Cost-related responses were provided under payer designations of self, third-party insurance, and federal government. RESULTS: The response rate was 81% (101/125 patients). tPA was preferred by 84%, and SK by 66%, for M alone and SR alone, respectively (chi 2, p < 0.01). Overall preference (M + SR) favored tPA (78%, p < 0.01). tPA preference decreased to 43% considering drug acquisition costs under the self-pay option (p < 0.01 vs M + SR). Similar trends of lesser magnitude were also observed for the third-party and government-payer options. CONCLUSIONS: Under conditions of zero cost and consideration of mortality plus stroke-risk data, tPA were preferred overall due to its lower mortality. Introduction of drug-cost data significantly shifted the preference toward SK, particularly under the self-payer designation. Patient preferences for thrombolytic therapy in AMI indicate tradeoffs between clinical attributes and costs, and should assist in framing medical debate and decision making.

Acute Disease↗
PubMedDate not supplied

Cilostazol.

Cilostazol is a phosphodiesterase III inhibitor that has been shown to be effective for treatment of symptomatic intermittent claudication. It is generally well tolerated, but is contraindicated in patients with severe heart failure because of the potential detrimental effects demonstrated by other agents in the same class. Additional studies are required to examine the long-term benefits of cilostazol therapy.

Cilostazol↗
PubMedDate not supplied

Cytochrome p450-mediated cardiovascular drug interactions.

Numerous cardiovascular agents are substrates, inhibitors, or inducers of cytochrome P450 (CYP450) enzymes. The growing understanding of the CYP450 enzyme family allows us to predict potential drug interactions. This review discusses CYP450-mediated cardiovascular drug interactions with selected examples reported in the literature. The principles of predicting drug interactions are also discussed. Studies evaluating in vivo drug interaction and case reporting by clinicians are necessary to establish the clinical significance of these interactions, which will in turn help to develop strategies to prevent and manage the potentially devastating effects they might have on patients.

Cardiovascular Agents↗
PubMedDate not supplied

Is the volume of distribution of digoxin reduced in patients with renal dysfunction? Determining digoxin pharmacokinetics by fluorescence polarization immunoassay.

STUDY OBJECTIVE: To determine digoxin pharmacokinetics in subjects with different degrees of renal function using fluorescence polarization immunoassay (FPIA), which is associated with less interference from digoxin-like immunoreactive substances (DLIS) than radioimmunoassay. SETTING: University hospital clinical research center. PARTICIPANTS: Eighteen subjects (mean age 44 yrs) with different degrees of renal function: group 1, creatinine clearance (Clcr) below 10 ml/minute; group 2, Clcr 10-50 ml/minute; and group 3, Clcr greater than 50 ml/minute (6 patients in each group). INTERVENTION: Over 5-7 days, 15 serum samples were collected after a single intravenous dose of digoxin 7 or 10 micrograms/kg actual body weight (WT) for serum concentration measurements by FPIA. Two-compartment pharmacokinetic parameters (zero-time intercept of the concentration-time curve of the initial distribution phase [A], zero-time intercept of the concentration-time curve of the terminal elimination phase [B], initial distribution phase constant [alpha], terminal elimination rate constant [beta], volume of distribution in the central compartment [Vc] and at steady state [Vss], total body clearance [Cl], mean residence time [MRT], area under the concentration-time curve [AUC]) were determined using a nonlinear least squares regression program. MEASUREMENTS AND MAIN RESULTS: No significant differences were found among groups for A, B, alpha, beta, beta-half-life Vc/WT, MRT, AUC, and Cl/WT. Significant differences were observed in Vss/WT (4.8 +/- 1.0, 6.6 +/- 0.5, 6.4 +/- 0.7 L/kg) between group 1 versus group 2 and group 1 versus group 3 (p < 0.01). Measured Clcr was correlated with Cl (r2 = 0.40, p < 0.01), Cl/WT (r2 = 0.29, p < 0.05), Vss (r2 = 0.35, p = 0.01), and Vss/WT (r2 = 0.24, p < 0.05). CONCLUSION: This study confirmed that Vss is smaller in patients with chronic renal failure (Clcr < 10 ml/min) than those without chronic renal failure. Therefore, previous recommendations that lower digoxin loading doses should be administered in patients with renal failure are applicable to digoxin serum concentration monitoring using FPIA.

Adolescent↗