Search PubMed⌕ Search

Biomedical subjects

L Ye

Publications and source records attributed to L Ye.

At least 109 records · Page 6Linked to original sources
PubMed1995

A microscopic model of enzyme kinetics.

Many in vivo enzymatic processes, such as those of the tissue factor pathway of blood coagulation, occur in environments with facilitated substrate delivery or enzymes bound to cellular or lipid surfaces, which are quite different from the ideal fluid environment for which the Michaelis-Menten equation was derived. To describe the kinetics of such reactions, we propose a microscopic model that focuses on the kinetics of a single-enzyme molecule. This model provides the foundation for macroscopic models of the system kinetics of reactions occurring in both ideal and nonideal environments. For ideal reaction systems, the corresponding macroscopic models thus derived are consistent with the Michaelis-Menten equation. It is shown that the apparent Km is in fact a function of the mechanism of substrate delivery and should be interpreted as the substrate level at which the enzyme vacancy time equals the residence time of ES-complexes; it is suggested that our microscopic model parameters characterize more accurately an enzyme and its catalytic efficiency than does the classical Km. This model can also be incorporated into computer simulations of more complex reactions as an alternative to explicit analytical formulation of a macroscopic model.

Animals↗
PubMed1995

Surface-mediated enzymatic reactions: simulations of tissue factor activation of factor X on a lipid surface.

Blood coagulation proceeds via reactions in which zymogen coagulation factors are activated to proteases. An essential step is the activation of factor X by a complex of tissue factor and factor VIIa. This complex usually is studied using phospholipid vesicles into which tissue factor is inserted. Because factor X exists free in solution and bound to the lipid-surface, it is difficult to establish experimentally the kinetic contribution of surfaces. We therefore developed a stochastic model to simulate such reactions and generate initial velocity data from which Michaelis-Menten parameters are estimated. Simulated Km values decrease slightly when substrate binding to lipid is increased and by a factor of four when the rates of surface diffusion are increased to that of fluid phase-diffusion. Simulations with various size planar surfaces established an enzyme capture radius of 32-64 nm. Simulations with different modes of enzyme-substrate complex assembly show that if the true substrate is lipid-bound, under certain conditions, the true Kcat is not measured; rather, the product "leaving rate" from the complex is the rate-limiting step that is measured as substrate is taken to infinity. This model is applicable to any surface-bound enzyme reaction.

Animals↗
PubMed1995

Viral shedding and fecal IgA response after Norwalk virus infection.

Protection is not conferred by preexposure to Norwalk virus (NV). By use of an ELISA with baculovirus-expressed recombinant NV (rNV) capsid protein, the pattern of NV fecal shedding and the protective effect of rNV-specific fecal IgA (flgA) were investigated in volunteers who were repeatedly challenged with NV. After the first challenge, ill volunteers were significantly more likely than well volunteers to have NV antigen in their stool (P < .05). After challenge, antigen shedding was detected on days 1-13; ill volunteers shed the antigen longer (P = .02). A higher prechallenge rNV-specific flgA geometric mean titer was found in ill compared with well volunteers (P < .05) and in infected versus noninfected volunteers (P < .05). NV shedding was common after infection and was present up to 2 weeks after challenge. Preexisting rNV-specific flgA, like serum IgG, is not protective and may be a marker for symptomatic disease.

Antibodies, Viral↗
PubMed1995

IP3 generation increases rod outer segment phagocytosis by cultured Royal College of Surgeons retinal pigment epithelium.

PURPOSE: To measure outer segment phagocytosis in cultures of Royal College of Surgeons (RCS) rat retinal pigment epithelium (RPE) that have been treated with carbachol. Carbachol treatment of RCS RPE results in an increase in the second messenger inositol triphosphate, which mimics that observed in normal RPE after interaction with rod outer segments (ROS). METHODS: Cultures of RCS RPE were phagocytically challenged with isolated rat ROS for 2 hours. Carbachol (1 mM) was added to some cultures to stimulate inositol triphosphate synthesis, and incubation continued for 30 minutes at 37 degrees C. Inositol triphosphate concentration was measured by radioreceptor assay. Bound and ingested outer segments were quantified by double immunofluorescent staining. Ingestion of outer segment membranes was confirmed by electron microscopy and immunogold staining. RESULTS: Carbachol treatment was associated with a rapid and temporary increase in inositol triphosphate levels. Royal College of Surgeons rat RPE phagocytically challenged with outer segments and treated with carbachol showed significantly higher ingestion (34%) compared to untreated RCS RPE (9%) (P < 0.05). CONCLUSIONS: Exposure of cultured RCS RPE to carbachol increases the intracellular concentration of inositol triphosphate and enhances phagocytosis of bound ROS. These results support the hypothesis that the phagocytic defect in RCS RPE is related to an abnormality in the generation of inositol triphosphate as a second messenger after outer segment recognition and binding.

Animals↗
PubMed1994

Design, characterization, and one-point in vivo calibration of a subcutaneously implanted glucose electrode.

A 0.29-mm-diameter flexible electrode designed for subcutaneous in vivo amperometric monitoring of glucose is described. The electrode was designed to allow "one-point" in vivo calibration, i.e., to have zero output current at zero glucose concentration, even in the presence of other electroreactive species of serum or blood. A valid zero point, along with a measurement of the glucose concentration in a withdrawn sample of blood at which the current is known, defined the sensitivity in the linear response range. The electrode was four-layered, with the layers serially deposited within a 0.125-mm recess upon the tip of a polyimide-insulated 0.25-mm gold wire. The recessed structure reduced the sensitivity to movement and allowed, through control of the depth of the recess, control of the transport of glucose and thus the range of linearity. The recess contained the four polymeric layers, with a total mass less than 5 micrograms and no leachable components. The bottom glucose concentration-to-current transducing layer consisted of the enzyme "wiring" redox polymer poly[(vinylimidazole)Os(bipyridine)2Cl]+ , complexed with recombinant glucose oxidase and cross-linked with poly(ethylene glycol) diglycidyl ether, to form an electron-conducting hydrogel. The layer was overcoated with an electrically insulating layer of polyaziridine-cross-linked poly(allylamine), on which an immobilized interference-eliminating horseradish peroxidase based film was deposited. An outer biocompatible layer was formed by photo-cross-linking derivatized poly(ethylene oxide). The current output of a typical electrode at 10 mM glucose and at 37 degrees C was 35 nA, the apparent Km was 20 mM, and the 10-90% response time was approximately 1 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Fragile X founder chromosome effects: linkage disequilibrium or microsatellite heterogeneity?

Previous studies of founder chromosome effects in fragile X have been based on linkage disequilibrium with either FRAXAC1 or DXS548 alone or combined with FRAXAC2. Recently, we found no linkage disequilibrium of FMR-1 with FRAXAC2, but rather, found FRAXAC2 was complex and highly mutable. Therefore, we have now analyzed FRAXAC1 and DXS548 together for haplotypes, two markers which have not been jointly analyzed previously, to test for disequilibrium. We typed 315 fragile X (FX) chromosomes and controls, further subdivided into large controls (LC) and small controls (SC) with < or = 35 repeats and identified 26 different haplotypes. Two were more frequent and one less frequent in FX than SCs, thus confirming apparent linkage disequilibrium in fragile X. However, we noted increased FX microsatellite heterozygosity, either individually (results quite similar to previous studies) or as haplotypes. This heterozygosity covaried with FX > LC > SC, which may indicate alternative explanation exists for the apparent disequilibrium. We hypothesize that large FMR-1 CGG repeat allele genes may be associated with the generation of new microsatellite mutations. Possible mechanisms include gene conversions between CGG repeats and flanking microsatellites involving unequal double cross-overs, the expansion of small control CGGs to larger sizes associated with episodic generalized microsatellite instability or as a direct result of mutant FMR-1 gene function. We conclude that the founder effects observed with the use of these CA repeats is likely to reflect both linkage disequilibrium and increased microsatellite instability of fragile X chromosomes.

Alleles↗
PubMed1994

Carrier detection of Werner's syndrome using a microsatellite that exhibits linkage disequilibrium with the Werner's syndrome locus.

Werner's syndrome (WS) is a rare autosomal recessive disorder, one of the progeroid syndromes, characterized by features of premature aging. The genetic defect in WS is unknown but recently the genetic linkage of WS to several markers on the short arm of chromosome 8 has been reported. Genetic analysis of 25 families with WS demonstrated that D8S339 was the closest marker linked to the gene locus for Werner's syndrome (WRN), with a peak lod score of 18.29 at recombination frequency 0.001, and showed a linkage disequilibrium with the WRN locus. We studied two unrelated families with WS using ANK1, D8S339, and D8S360. The mutative haplotype identified through the generations in pedigrees provides a means of carrier detection and presymptomatic diagnosis.

Female↗
PubMed1994

[Determination of salbutamol in human urine].

Salbutamol (2.4 mg) was administrated to a healthy male. The urine was analysed by GC-MS. 95% of the drug was excreted in 24 h in unconjugated form. The pH of urine was shown to strongly affect the extraction rate, good recovery was obtained at pH 8.8. With this method, salbutamol can be determined simply, quickly and accurately.

Albuterol↗
PubMed1994

[Trenbolone and its metabolites in human urine by GC/MS analysis].

The gas chromatography/mass spectrometric properties of trimethylsilyl ether and trifluoroacetyl ester derivatives of trenbolone, combined with a methoxine group, are presented. The cleavage mechanism and some derivatization problems were observed and discussed. The positive urine of trenbolone acetate was checked with the results of epitrenbolone and possible hydroxy-trenbolone as main metabolites. Also a little trenbolone and possible estra-4,9,11-trien-3,17-dione were found.

Anabolic Agents↗
PubMed1993

Isolation and mapping of microsatellites from a library microdissected from the Werner syndrome region, 8p11.2-p22.

We have constructed a new genetic linkage map of the Werner syndrome (WRN) region, using microsatellites from a library which was developed by a chromosome microdissection and enzymatic amplification method. These microsatellites were used to genotype members of CEPH families using a simplified detection system of polymerase chain reaction (PCR) products. Two-point analysis was used to assign 4 microsatellite markers relative to each marker and other markers reported in the CEPH public data base. We confirmed that these 4 markers are located to the WRN region, 8p11.2-p22. Such microsatellites microdissected from the definite chromosome region may be useful for positional cloning.

Base Sequence↗