Biomedical subjects

N Yuan

Publications and source records attributed to N Yuan.

9 recordsLinked to original sources

PubMed1996

Multichannel flow electrophoresis and its applications in purification of proteins, enzymes, and antibodies.

Multichannel flow electrophoresis (MFE) is a novel preparative electrophoresis technique designed for continuous separation of proteins based on their different isoelectric points. A computer controlled MFE apparatus is developed and the effects of operation parameters such as electric field strength, buffering pH, and sample introducing flow rate on MFE separation efficiency are investigated. The application of MFE is first exemplified by continuous separation of BSA and HBB mixture. Then continuous purification of urokinase from urine extracts is conducted, in which 10 x 10(4) IU urokinase product whose specific activity is higher than 40,000 IU/mg is obtained per hour. Finally, MFE is applied in the continuous purification of anti-urokinase mouse IgG from mouse serum and yields 2.1 mg mouse IgG per hour with high purity demonstrated by SDS-PAGE. The preliminary results presented have confirmed the workability of MFE and pointed out its high application potential in biochemical process.

Antibodies↗

PubMed1994

cAMP-dependent protein kinase and protein kinase C consensus site mutations of the beta-adrenergic receptor. Effect on desensitization and stimulation of adenylylcyclase.

Activation of cAMP-dependent protein kinase (cAPK) or protein kinase C (PKC) causes a rapid desensitization of beta 2-adrenergic receptor (beta AR) stimulation of adenylylcyclase in L cells, which previous studies suggest involves the cAPK/PKC consensus phosphorylation site in the third intracellular loop of the beta AR, RRSSK263. To determine the role of the individual serines in the cAPK- and PKC-mediated desensitizations, wild type (WT) and mutant beta ARs containing the substitutions, Ser261-->Ala, Ser262-->Ala, Ser262-->Asp, and Ser261/262-->Ala, were constructed and stably transfected into L cells. Results showed that serine 262 was the primary site of the cAPK-induced desensitization, whereas either serine 261 or serine 262 was sufficient to confer the 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA)/PKC-mediated desensitization. Coincident stimulation of cAPK and PKC caused an additive desensitization (6-8-fold increase in the EC50) which was significantly reduced (80%) only by the double substitution mutation. Quantitative evaluation of the coupling efficiencies and the GTP-shift of the WT and mutant receptors demonstrated that only one of the mutants, Ser262-->Ala, was partially uncoupled. The Ser262-->Asp mutation did not significantly uncouple, demonstrating that introducing a negative charge did not appear to mimic the desensitized state of the receptor. The beta AR expression level played a critical role in determining the pattern of beta AR desensitization; i.e. while the overall desensitization was unaltered within a large range of beta AR expression level (10-300 fmol/mg), the increase in EC50 and decrease in Vmax were differentially affected by the change in the receptor level.

Adenylyl Cyclases↗

PubMed1994

Pseudohomogeneous kinetic study on a two-liquid-phase fermentation process.

The fermentation process for producing undecane dicarboxylic acid from tridecane, which includes gas-oil-water-cell four phases (two-liquid-phase), was studied. The metabolic characters of the cell growth phase and the production phase of the process were analyzed. It was proposed that cell growth can be identified by the carbon dioxide production rate (CPR) before the production phase. The kinetic models of both the cell growth phase and the production phase were established, respectively. The parameters of the models have been estimated by regression. The calculated curves fit the experimental data very well. The average deviation between those over the cell growth phase and the production phase are 2.4% and 3.6%, respectively.

Alkanes↗

PubMed1994

Differential expression of the beta-adrenergic receptor modifies agonist stimulation of adenylyl cyclase: a quantitative evaluation.

The effect of varying levels of beta 2-adrenergic receptor (beta AR) expression on the capacity of the receptor to activate adenylyl cyclase through regulatory G proteins has been systematically explored in this paper, using differential expression of hamster and human beta AR in L cells. Equations have been developed based on the cycle of G protein activation first proposed by Cassel and Selinger and the mobile receptor model, which assumes that hormone-bound beta AR can stimulate a number of different adenylyl cyclase moieties through the G protein during a single cycle of activation. These equations predict the relationship of receptor number to the EC50 (potency) and Vmax (efficacy) for adenylyl cyclase activation. L cell clones were selected with stable expression of the beta AR over a 2000-fold range of levels (from 5 to 10,000 fmol/mg of membrane protein). Experimentally determined values for the EC50 and the Vmax for epinephrine stimulation of adenylyl cyclase over the entire range of receptor levels were found to be in excellent agreement with predictions of the traditionally accepted models. A method is introduced that allows calculation of beta AR coupling efficiency while taking into account the effect of variable receptor levels. The approach provides a quantitative means for the determination of coupling efficiency of the receptor/G protein/adenylyl cyclase system over wide variations in receptor levels and allows for a rational comparison of coupling efficiencies of wild-type and mutant receptors when receptor levels differ.

Adenylyl Cyclases↗

PubMed1992

Regulation of the renal Na(+)-H+ exchanger by calcium calmodulin-dependent multifunctional protein kinase II.

Prior studies indicate that cAMP-dependent protein kinase (PKA) and calcium calmodulin-dependent multifunctional protein kinase II (CaM-KII) inhibit Na(+)-H+ exchanger as assayed in octyl glucoside solubilized rabbit renal brush border membrane proteins reconstituted into artificial lipid vesicles. An anion exchange chromatography fraction of these proteins which elutes between 0.2 and 0.4 M NaCl (Fraction B), however, fails to demonstrate regulation of the transporter by PKA. The present studies examine regulation of the Na(+)-H+ exchanger by CaM-KII using Fraction B proteins. As compared to the initial total protein extract, Fraction B demonstrated increased Na(+)-H+ exchange activity. CaM-KII inhibited the Na(+)-H+ exchanger in Fraction B by 38.2 +/- 10.6% in an ATP and calmodulin-dependent manner. The results of the present studies suggest that CaM-KII-mediated inhibition of the Na(+)-H+ exchanger involves the phosphorylation of different polypeptides than those mediating the inhibition of this transporter by PKA.

Adenosine Triphosphate↗

PubMed1991

The influence of adaptation on the oscillatory potentials of the human electroretinogram.

The influence of adaptation on the oscillatory potentials of the human electroretinogram was studied in the domains of frequency and time. The amplitude of OP1 to OP4, the summed amplitude of OP1 to OP4, as well as the area, decreased from dark adaptation to light adaptation. With increasing intensities of background illumination, they increased slightly and appeared to decrease with the strongest background illumination. The implicit time of oscillatory potentials 3 and 4 increased with stronger background illumination and decreased with the strongest background illumination. The results of the dominant frequency and the total power of the OPs correspond to the results in the time domain. The dominant power decreased from dark adaptation into light adaptation and did not show any systematic changes with increasing intensity of background illumination.

Adult↗

PubMed1991

Residual H2O2 compromises deswelling function of in vivo rabbit cornea.

The purpose of the study was to investigate the effect of H2O2 on corneal swelling, corneal deswelling and to determine the concentration of H2O2 in the aqueous humor. Four types of experiments were performed using the rabbit eye. 1. Central corneal thickness (CCT) measurements after instillation of H2O2 into the cul-de-sac and after wearing H2O2 soaked soft contact lenses (SLC) for 2 h using 60 ppm, 100 ppm and 300 ppm H2O2. 2. Measurement of corneal deswelling induced by wearing SCL and closed eye for 2 h. 3. Determining the H2O2 in the aqueous humor one h after instillation of H2O2 into the cul-de-sac. CCT increases of 7.5% and 10.7% were found after the instillation of 100 ppm and 300 ppm H2O2 into the cul-de-sac. After wearing H2O2 soaked SCLs, CCT increased 4.6% and 5.8%, respectively. The H2O2 in the aqueous humor was 0.087 mM or a 160% increase after using 300 ppm H2O2. The increase in CCT and interference with the deswelling process is interpreted as a toxic effect on the cornea and an interference with the normal dehydration function of the endothelial pump.

Administration, Topical↗

PubMed1990

Measurement of the oscillatory potential of the electroretinogram in the domains of frequency and time.

The dark-adapted and light-adapted electroretinograms of 13 subjects with 23 normal eyes were analyzed by means of Fourier spectrum. The oscillatory potentials in the time domain were filtered out from the electroretinogram after a corresponding bandpass was given in the frequency domain. The coefficient of variation of total power, dominant power and dominant frequency of the isolated oscillatory potentials in the frequency domain, summed amplitudes and area of the isolated oscillatory potentials, each amplitude and implicit time of the first four major oscillatory potential wavelets in the time domain were compared. The implicit time showed the smallest coefficient of variation; summed amplitudes of OP1 to OP4 showed smaller coefficients of variation than those of the area, the amplitude of each oscillatory potential wavelet, dominant frequency and dominant and total power. The coefficient of variation of these measurement parameters in light-adapted electroretinograms was smaller than those in dark-adapted electroretinograms.

Adult↗

PubMed1990

Regulation of renal Na(+)-H+ exchanger by cAMP-dependent protein kinase.

Octyl glucoside-extracted rabbit renal brush-border membrane (BBM) proteins were sequentially fractionated using anion exchange chromatography, and the fractions were tested for Na(+)-H+ exchange activity, amiloride sensitivity, and the effect of adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase (PKA) after reconstitution into artificial lipid vesicles. Compared with the initial protein extract, an anionic protein fraction eluting with 0.2-0.4 M NaCl (fraction B) demonstrated increased Na(+)-H+ exchange activity. Fraction B also demonstrated sensitivity to inhibition by amiloride but was not regulated by PKA. Co-reconstitution of fraction B with a BBM protein fraction highly enriched in a 42-kDa polypeptide restored the inhibitory response to PKA. These experiments suggest that, as assayed in a solubilized and reconstituted system, the Na(+)-H+ exchanger contains a dissociable PKA regulatory component, possibly a polypeptide of 42 kDa.

Amiloride↗