Summary Four antifreeze proteins (AFPs) were purified from larvae of the beetle Dendroides canadensis. The AFPs are similar in amino acid compositions, having high contents of hydrophilic amino acids (45–55 mol%) and cysteine (16 mol% Cys). Approximately half of the Cys residues form disulfide bridges, and both the disulfide bridges and free sulfhydryls are essential for activity. The N-terminals of the AFPs are blocked. The pH optimum of the AFPs is 7.8, but major loss of activity occurred only at very high pH (12.0). The detergents SDS and Triton X-100 did not inactivate the AFPs. Circular dichroism spectra indicate the presence of both and secondary structures in the AFPs, in addition to a large random structure component.Abbreviations
AFP
antifreeze protein
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CD
circular dichroism
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DTT
dithiothreitol
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HPLC
high pressure liquid chromatography
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PAGE
polyacrylamide gel electrophoresis
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PAS
periodic acid Schiff
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SDS
sodium dodecyl sulfate
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TFA
trifluoroacetic acid 相似文献
We have further characterized the 5-HT3 receptors in rat and rabbit tissues by evaluating the binding of the 5-HT3 receptor ligand, [3H]GR67330 to homogenates of rabbit ileum, rat ileum and rat brain (entorhinal cortex). In each tissue specific [3H]GR67330 binding represented a single saturable, high affinity site (Kd = 0.14, 0.18, 0.076 nM in rabbit ileum, rat ileum and rat brain respectively). The densities of sites present in rabbit and rat ileum were similar to that present in rat brain (Bmax = 63, 47, 72 fmol/mg protein in rabbit ileum, rat ileum and rat brain respectively).
In each tissue, 5-HT3 receptor agonists and antagonists potently competed for [3H]GR67330 binding. Derived inhibition constants were similar in rat ileum and brain. However marked differences in IC50s were apparent for rabbit ileum compared with rat brain or ileum. These were most apparent with agonists. Thus, mCPBG [1-(meta-chlorophenylbiguanide)], phenylbiguanide, 5-HT and 2-methyl 5-HT were at least 5 times less potent to inhibit [3H]GR67330 binding in rabbit ileum than rat brain. The most pronounced differences were evident with phenylbiguanide and mCPBG which were 70 and 300 times less potent in the rabbit ileum respectively compared with the rat tissues. These differences were unlikely to be due to depletion effects because tissue combination experiments (rabbit ileum and rat brain) yielded biphasic inhibition curves for phenylbiguanide with affinities for each component similar to those in the individual tissues. Antagonist affinities also varied between the rabbit and rat tissues, although less markedly. Amongst the antagonists, the most marked differences were apparent with SDZ 206–830 and quipazine each being 10 times less potent to inhibit binding to rabbit than rat tissue.
Hill coefficients for inhibition of binding varied with tissue. In rat brain, as previously described for [3H]GR67330, Hill coefficients for agonist (and quipazine) inhibition of binding were greater than unity. This was less marked in rat and rabbit ileum tissues.
The present studies provide further evidence for species variation in 5-HT3 receptors. 相似文献
The cardiovascular responses to a 10-min 1.22 rad (70 degrees) head-up tilt orthostatic tolerance test (OST) was observed in eight healthy men following each of a 5-min supine baseline (control), 4 h of 0.1 rad (6 degrees) head-down tilt (HDT), or 4 h 0.52 rad (30 degrees) head-up tilt (HUT). An important clinical observation was presyncopal symptoms in six of eight subjects following 4 h HDT, but in no subjects following 4 h HUT. Immediately prior to the OST, there were no differences in heart rate, stroke volume, cardiac output, mean arterial pressure and total peripheral resistance for HDT and HUT. However, stroke volume and cardiac output were greater for the control group. Mean arterial pressure for the control group was less than HDT but not HUT. Over the full 10-min period of OST, the mean arterial pressure was not different between groups. Heart rate increased to the same level for all three treatments. Stroke volume decreased across the full time period for control and HDT, but only at 3 and 9 min for HUT. There was a higher total peripheral resistance in the HDT group than control or HUT. The pre-ejection period to left ventricular ejection time ratio was less in HDT than for control or HUT groups. These data indicate a rapid adaptation of the cardiovascular system to 4 h HDT that appears to be inappropriate on reapplication of a head to foot gravity vector. We speculate that the cause of the impaired orthostatic tolerance is decreased tone in venous capacitance vessels so that venous return is inadequate. 相似文献
A human subtelomeric repeat (designated as the HST repeat) has been isolated and characterized from a yeast artificial chromosome containing one human telomere. This repeat is located immediately adjacent to the telomeric T2AG3 repeats at the extreme termini of the human chromosomes. The DNA sequence of 3.6 kb of the HST repeat has been determined. The HST repeat spans over 3.6 kb in length, and contains one evolutionarily conserved CpG-rich region. The copy number of the HST repeat varies among telomeres. Genomic hybridization experiments suggest that the HST repeat consists of two distinct segments, and the distal portions of the HST repeat are also distributed elsewhere in the genome. In HeLa cells, the HST repeat sequence appears to be transcribed into a 6 kb polyadenylated RNA and a variety of non-polyadenylated RNA species. 相似文献
Isolated ovarian corpus luteal cells and granulosa cells of rat were employed to investigate the effect of alpha-ANP on the secretion of progesterone and estradiol. The contents of the steroid hormones are determined by RIA. The results showed that 0.1-10 ng/ml ANP promoted progesterone production in a dose dependent manner. alpha-ANP also enhanced progesterone production by granulosa cells, but not estradiol. It seems that the effect of alpha-ANP on ovarian steroidogenesis is a direct one. 相似文献