Assaying proteins for molecular chaperone activity.
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Animals↗
Biomedical subjects
G J Lee
Publications and source records attributed to G J Lee.
Leucine 332 influences the CO2/O2 specificity factor of ribulose-1,5-bisphosphate carboxylase/oxygenase from Anacystis nidulans.
The role of Leu 332 in ribulose-1,5-bisphosphate carboxylase/oxygenase from the cyanobacterium Anacystis nidulans was investigated by site-directed mutagenesis. Substitutions of this residue with Met, Ile, Val, Thr, or Ala decreased the CO2/O2 specificity factor by as much as 67% and 96% for the Ile mutant in the presence of Mg2+ and Mn2+, respectively. For the Met, Ile, and Ala mutants in the presence of Mg2+, no loss of oxygenase activity was observed despite the loss of greater than 65% of the carboxylase activity relative to the wild-type enzyme. In the presence of Mn2+, carboxylase activities for mutant enzymes were reduced to approximately the same degree as was observed in the presence of Mg2+, although oxygenase activities were also reduced to similar extents as carboxylase activities. Only minor changes in Km(RuBP) were observed for all mutants in the presence of Mg2+ relative to the wild-type enzyme, indicating that Leu 332 does not function in RuBP binding. These results suggest that in the presence of Mg2+, Leu 332 contributes to the stabilization of the transition state for the carboxylase reaction, and demonstrate that it is possible to affect only one of the activities of this bifunctional enzyme.
Carbon Dioxide↗
Genetic basis of prolificacy in Meishan pigs.
Research in France and in the UK confirms the prolificacy of the Chinese Meishan breed to be about three to four piglets greater than that of control Large White females. Crossbreeding studies clearly indicate that this breed difference is due to genes acting in the dam and not in the litter itself. There is high heterosis for litter size in F1 Meishan x Large White crossbred females, such that their litter size is similar to or greater than that of purebred Meishan females. There is some discrepancy between studies about whether the Meishan female has a higher ovulation rate than does the Large White breed and this can be attributed in large part to the different basis upon which breed comparisons have been made. Nevertheless, there may be real genetic differences between Meishan pigs exported to different countries. In young gilts at comparable numbers of oestrous cycles after puberty, the ovulation rate is similar in Meishan and Large White gilts, but in older gilts, and particularly in multiparous sows, Meishan pigs have a higher ovulation rate in British studies. Once comparisons of prenatal survival between breeds have been adjusted for any breed difference in ovulation rate, the main cause of prolificacy in Meishan pigs can be seen to be an enhanced level of prenatal survival. Crossbreeding studies show that this is controlled by the maternal genotype and not that of the embryos. The advantage in prenatal survival to the Meishan pig is clearly present in the post-attachment period (after day 20 of gestation), but may also be present earlier in gestation. Results from a study presented here suggest that Meishan sows have a higher uterine capacity than do Large White sows and this allows them to maintain their higher number of attached embryos through gestation. F1 Meishan x Large White crossbred females achieve their high litter size via a different route than do purebred Meishan females. These animals have a lower ovulation rate and fewer attached embryos than do purebred Meishan sows, but a very low level of fetal loss allows them to produce litters of similar size. The low level of fetal loss in F1 females appears to be due to the higher uterine capacity of F1 females compared with purebred Meishan sows.
Animals↗
Serine-376 contributes to the binding of substrate by ribulose-bisphosphate carboxylase/oxygenase from Anacystis nidulans.
Site-directed mutagenesis was used to change Ser376 in the active site of ribulose-1,5-bisphosphate carboxylase/oxygenase from the cyanobacterium Anacystis nidulans to Cys, Thr, or Ala. When expressed in Escherichia coli and purified, the mutant enzymes exhibited carboxylase activities that were reduced by 99% or more with respect to the activity of the wild-type enzyme. The Km values for ribulose bisphosphate at pH 8.0, 30 degrees C, were elevated from 46 microM for wild-type enzyme to 287, 978, and 81 microM for mutants in which Cys, Thr, or Ala, respectively, replaced Ser376. The Cys and Thr variants were almost devoid of oxygenase activity whereas the Ala variant had 16% as much oxygenase as wild-type enzyme, suggesting that this mutation had greatly elevated the oxygenase:carboxylase ratio.
Alanine↗
The budgetary impact of 5-HT3 receptor antagonists in the management of chemotherapy-induced emesis.
The study examined the budgetary implications of using 5-hydroxytryptamine3 receptor antagonists (5-HT3RA), granisetron or ondansetron, in the management of chemotherapy-induced emesis (CIE). A treatment model was constructed to represent a baseline of efficacy and costs for treating a cohort of patients with conventional antiemetics. Groups of patients who would be expected to receive the most benefit from 5-HT3RA were then identified and the effect upon costs of using these compounds in a consecutively larger proportion of selected patients was calculated. On the basis of illustrative costs from The Cookridge Hospital in the UK, it was concluded that the new antiemetics can be used in acute emesis with substantial clinical benefit for an increase of 3-10% to total treatment costs. However, for delayed emesis these compounds have not yet shown a clinical advantage, and the increase in total costs of 12-34% is not justified.
Antiemetics↗
Direct responses in males and correlated responses for reproduction in females to selection for testicular size adjusted for body weight in young male lambs.
Selection based upon testicular diameter adjusted for body weight at 6, 10 and 14 weeks of age was used to produce two lines of sheep, with either high or low testicular size. Ten generations of selection were carried out and the estimate of the realized heritability of the selection criterion was 0.53 +/- 0.01. There were significant positive correlated responses to selection for testicular diameter at 6, 10 and 14 weeks of age, but the correlated responses in body weight at these ages were negative. In mature females, there were significant negative correlated responses to selection in premating body weight in the 1st, 2nd and 3rd breeding season and in the day of the first oestrus in the 2nd breeding season. Litter size per ewe mated had a small positive correlated response to selection in the second breeding season. This latter response appeared to be due to a positive correlated response in fertility, ewes from the High-line having a significantly higher probability of conceiving to a single mating than those from the Low-line. There was no significant correlated response in ovulation rate or litter size per ewe lambing and the genetic correlation between these traits and the selection criterion is likely to be close to zero. This may be due to the adjustment for body weight used, but it is possible that, in any event, body weight in young rams may be a better predictor of female ovulation rate than testicular diameter. These results do not rule out the possibility that testicular size in rams older than those selected would provide a good predictor of genetic merit for female ovulation rate.
Animals↗
Pharmacokinetics of norethindrone: effect of particle size.
In 24 healthy women between the ages of 19 and 35 years who had not used oral contraceptive preparations for at least 60 days, it was found that the smaller the particle size of norethindrone (NET) administered, the higher was the plasma NET level obtained. Three different preparations having particle sizes of NET smaller than 250 microns, 44 microns or 10 microns were tested in a crossover pattern. The time required to reach maximum plasma concentration (Tmax) became shorter with decreasing particle size, 1.69 hr, 1.52 hr and 1.06 hr, respectively. As particle size was reduced, the maximum NET plasma concentration (Cmax) increased for the 3 different 1 mg NET preparations, i.e. 8.66 ng/ml, 10.53 ng/ml and 15.73 ng/ml. A trial with a 2 mg NET preparation made with NET utilizing the 44 microns same material displayed a Tmax similar to the 1 mg NET preparation having the same particle size while the Cmax reached a level of 17.56 ng/ml. The area under the plasma concentration versus time curve from 0-24 hrs and the extrapolated total area under the curve, increased with decreasing particle size. The use of a smaller particle size allows for more rapid dissolution or oral contraceptive tablets when measured in vitro; however, there is no evidence that such faster dissolution leads to a significant difference in efficacy. Oral contraceptive tablets have, since their inception, utilized both large and small NET particle size material in various preparations.
Administration, Oral↗
Bioequivalence of two oral contraceptive drugs containing norethindrone and ethinyl estradiol.
Two oral contraceptive drugs, Formulation A and Formulation B, both of similar hormonal content, were compared with each other to determine if they were bioequivalent. Both drugs contain 1 mg of norethindrone (NET) and 0.035 mg of ethinyl estradiol (EE). Application of an interval test for the ratio of the computed parameter means demonstrated equivalence for the two formulations with respect to the 0-24 hour area under the plasma level versus time curve (AUC24), the total area under the curve (AUCtot) and for the maximum plasma concentration (Cmax) for both ethinyl estradiol and norethindrone. The data support the hypothesis for bioequivalence of the two formulations with respect to total absorption.
Adult↗
Study of LH response to GnRH in the young male as a criterion of genetic merit for female reproduction in sheep.
A high and a low response line in sheep were selected on the basis of the mean concentration of LH in 10-week-old Finn-Dorset ram lambs after an i.v. injection of 5 micrograms GnRH. After 8 male generations the mean LH response of the high line was more than 5-fold that of the low line and the heritability of the selected trait was estimated at 0.44 +/- 0.015. Highly significant line differences in mean LH response to GnRH were also found in males at 20 weeks of age and females at 10 and 20 weeks of age and the genetic correlations between the four LH response traits appear to be close to unity. Large line differences in the mean FSH response to GnRH were also found in both males and females at 10 and 20 weeks of age. Selection had little effect on the physical characteristics of lambs. High-response line ewes entering their first breeding season at about 7 months of age showed oestrus earlier in the season and had higher ovulation rates and numbers of lambs born per ewe lambing than did low-response line ewes. In the second breeding season, at about 19 months of age, the only line difference was a higher ovulation rate early in the breeding season in high-line ewes. It is suggested that these changes may be mediated by a more rapid response in high-line ewes to increased GnRH stimulation at puberty or at the beginning of the breeding season.
Animals↗
[The effect of muscle relaxation training on patients with insomnia complaints].
This study was performed to examine the effect of muscle relaxation training on patients with insomnia complaints. The subjects were selected those who have taken poor sleep below 240-300 minutes per day, between 16-15 ages, without organic brain syndromes. Using the Budzinski's muscle relaxation training tapes the patients have been gotten the muscle relaxation training daily during four weeks and examined the state anxiety with Spielberger's Trait-State Anxiety Scale, and investigated Sleep amounts, Blood pressure, Pulse rates, Respiratory rates before and after the muscle relaxation training. The results were as follow: 1. The mean of the State anxiety after the muscle relaxation training (41.8 +/- 6.4) was significantly lower than that of before training (54.2 +/- 7.0) (p less than 0.001). 2. The mean of systolic blood pressure after the training (114.3 +/- 7.8 mmHg) was significantly lower than that of before training (139.0 +/- 9.8) (p less than 0.001). 3. The mean of Pulse rate after the training (89.2 +/- 3.0) was significantly lower than that of before training (103.9 +/- 7.4) (p less than 0.001). 4. The mean of Respiratory rate after the training (18.6 +/- 1.0) was significantly lower than that of before training (22.8 +/- 1.3) (p less than 0.001). 5. The mean of Sleep amount after the muscle relaxation training (459.8 +/- 52.4 minutes) was significantly increased than that of before training (287.3 +/- 30.3) (p less than 0.001).
Anxiety↗
Isolation of temperature-sensitive tyrosine kinase mutants of v-abl oncogene by screening with antibodies for phosphotyrosine.
Temperature-sensitive protein-tyrosine kinase (EC 2.7.1.112) mutants of the oncogene v-abl have been obtained by a direct screening of kinase mutants in bacteria. The v-abl oncogene was expressed in Escherichia coli as a trpE/v-abl fusion protein from the trp promoter. The expression plasmid was mutagenized in vitro and then transfected into E. coli. Bacteria that produced defective tyrosine kinases were distinguished from those producing wild-type v-abl kinases by hybridization with antibodies specific for phosphotyrosine. Two independent mutations that generated temperature-sensitive tyrosine kinases were found to be located in a 12-amino acid region in the tyrosine kinase domain of the v-abl-encoded protein. These mutant v-abl oncogenes displayed temperature-sensitive transforming activity when expressed in NIH 3T3 cells. Cells transformed by these temperature-sensitive tyrosine kinase mutants could be shifted between the transformed and untransformed states by changing their growth temperature. These results confirmed the crucial role of tyrosine kinase activity in the v-abl-mediated oncogenesis.
Amino Acid Sequence↗
Bioequivalence of norethindrone and ethinyl estradiol for two different weight tablets with the same hormonal content.
Two tablets of differing weights, 100 mg (A) and 50 mg (B), of an oral contraceptive drug (OC) were compared with each other and to a solution (C) of the same components. The composition of the OC consisted of 1 mg of norethindrone (NET) and 0.035 mg of ethinyl estradiol (EE2). Both tablets were shown to differ from the solution, which was more rapidly absorbed, but were not significantly different from each other. Formulation means for NET and EE2, for each of the two products, were similar. Application of an interval test for the ratio of computed parameters demonstrated equivalence of the two formulations with respect to 0-24 hr area under the curve (AUC24) and total area under the curve (AUC+o+) for both NET and EE2 and with respect to concentration maximum (Cmax) for EE2. The data support the hypothesis for bioequivalence of the two formulations with respect to total absorption.
Absorption↗
Endocrine differences in rams after genetic selection for testis size.
Testis diameter and body weight were recorded from 6 to 76 weeks of age in ram lambs from two established lines selected for high (H) and low (L) testis size. While testis growth was greater in the H line up to 14 weeks of age (P less than 0.001), body weight was significantly lower, with the L line rams being 10 kg heavier by 76 weeks. There were no differences in plasma LH up to 20 weeks of age, but FSH concentrations were significantly lower at 14 and 20 weeks in the H line. Testosterone concentrations were not significantly higher in the H line from 6 to 20 weeks. In lambs castrated at birth, significantly higher FSH values were recorded from 6 to 20 weeks of age in the H line (P less than 0.001) whereas there was no difference in LH concentration at 6 and 10 weeks of age between the lines. At 14 and 20 weeks, however, the concentrations of LH were greater in the H than L line lambs (P less than 0.05). After hemicastration at 6 weeks of age, the rate of growth of the remaining testis in the L line lambs was significantly faster than in entire lambs of that line from 10 to 20 weeks (P less than 0.05 at 10 weeks to P less than 0.001 at 20 weeks). There was no difference in the rate of testis growth between the the entire and hemicastrated lambs from the H line from 6 to 12 weeks of age. It can be concluded that there is an underlying genetic difference in pituitary gland and/or hypothalamic activity in ram lambs from the two selected lines.
Animals↗
Culling and wastage in dairy cows in East Anglia.
A culling survey in 80 Friesian dairy herds in East Anglia over four years investigated the age at, and reasons for, disposal or death. Failure to conceive was the most important reason except in very old animals. Low production was the second most common reason, culling being particularly heavy in the first two lactations. Mastitis was the major disease influencing wastage and increased with age until the sixth lactation. The percentage of culls for multiple reasons increased with age. There was a high proportion of young animals in most herds and the median lactating life of cows was about three years, equivalent to three lactations. Considerable variation in herd life demonstrated that there was substantial scope for improving herd longevity. Long living herds culled fewer animals for breeding problems in the early lactations and for mastitis in the later ones, enabling more animals to be culled as surplus and for production factors.
Age Factors↗
Separation of reduced disaccharides derived from glycosaminoglycans by high-performance liquid chromatography.
A high-performance liquid chromatographic (HPLC) method for the analysis of reduced unsaturated disaccharides derived from enzymatic digestion followed by reduction with sodium borohydride of chondroitin sulfates, dermatan sulfate, heparan sulfate and heparin is described. This method is well suited for the HPLC analysis of glycosaminoglycans (GAGs) because the possibility of obtaining anomeric forms of unsaturated disaccharides is eliminated with provides a major advantage for quantitation. This procedure is more sensitive than existing HPLC methods for the determination of enzymatic degradation products from GAGs. In particular, the resolution of disaccharide products from heparan sulfate is improved after reduction. The applicability of this method for the determination of GAGs in biological samples is demonstrated.
Chondroitinases and Chondroitin Lyases↗
The application of high-performance liquid chromatography in enzymatic assays of chondroitin sulfate isomers in normal human urine.
A study of the urinary excretion of isomeric chondroitin sulfates in normal individuals by high-performance liquid chromatographic (HPLC) determinations of the unsaturated disaccharides produced by digestion with chondroitinases is described. The composition of the HPLC mobile phase was systematically varied in order to select the optimal conditions for separation. The data show that chondroitin 4-sulfate is the major component of the chondroitin sulfate isomers in normal urine, and that chondroitin 6-sulfate is a lesser component. It is also evident that dermatan sulfate is present in small quantities in normal urine.
Adult↗
Separate contribution of ventricular pumping and load impedance on ventricular power output.
The heart is a rather special type of fluid pump in the sense that it is an organic pump. In this paper, however, it is postulated that the heart behaves like any other mechanical pump whose power output may assume, for a primary change in load, values bounded by two limits which are power output values obtained had the pump been a pressure generator and a flow generator respectively. A power output value outside the limits would imply a definite alteration in ventricular pump function concomitant with the load change. A nomogram to illustrate the proposal is included. The hypothesis may be developed into a powerful tool for assessing the pumping capability of the left ventricle, and this has significant clinical and physiological implications.
Animals↗
Enzymatic studies of urinary isomeric chondroitin sulfates from patients with mucopolysaccharidoses. The application of high performance liquid chromatography.
The high-performance liquid chromatographic (HPLC) method for the determination of unsaturated sulfated disaccharides is a comprehensive and reliable method which expedites ensymatic studies of isomeric chondroitin sulfates. Responses for these unsaturated disaccharides derived from urinary chondroitin sulfates were linear from 100 ng to 10 micrograms injected and good quantitation was obtained for 25 microliters or less of samples placed on the column. This method which is more sensitive and accurate than methods now being used has considerable potential for the chemical diagnosis of patients with mucopolysaccharidoses and for the clarification of glycosaminoglycan structure. The isomeric chondroitin sulfates in urines from patients with mucopolysaccharidoses were studied by enzyme digestion with chondroitinases followed by HPLC determination of the sulfated unsaturated disaccharides produced. Evaluation by HPLC of the unsaturated 4-sulfated disaccharide produced by digestion of the urinary GAG with chondroitinases ABC and AC revealed rapidly and quantitatively the large amounts of dermatan sulfate present in Hurler, Hunter, and Maroteaux-Lamy urines. Chondroitin 4-sulfate predominated in Sanfilippo urinary isomeric chondroitin sulfates whereas chondroitin 6-sulfate and chondroitin 4-sulfate were shown to be present in nearly equal amounts in Morquio urine. An oversulfated chondroitin sulfate was detected in small amounts in some of these urines. This was demonstrated by the detection of an unsaturated disulfated disaccharide after digestion with chondroitinase ABC but not with chondroitinase AC.
Chondroitin↗