Analysis of teleost hemoglobin by Adair and Monod-Wyman-Changeux models

Summary The allosteric effects of the erythrocytic nucleoside triphosphates (NTP) and of proton concentrations were investigated by precise measurement of Hb–O₂ equilibria of tench hemoglobin (including extreme, high and low saturation ranges) and analysed in terms of the MWC two state model and the Adair four step oxygenation theory.At low concentrations (NTP/Hb ratio=1.0, and pH>7.3) ATP, GTP and protons decrease Hb–O₂ affinity by increasing the allosteric constantL and reducingK _T, the association constant¹ of the deoxy, tense state of the Hb, without significantly affecting that (K _R) of the oxy state, increasing the free energy of cooperativity (G). High concentrations of these effectors, however, also reduceK _R. The greater sensitivity of the half-saturation O₂ tension (P ₅₀) of the Hb to GTP than to ATP at the same concentration, correlates with greater effects of GTP on bothK _T andK _R. The pH and NTP dependence of the four Adair association constants and the calculated fractional populations of Hb molecules in different stages of oxygenation show that the autochthonous NTP effectors and protons stabilize the T structure and postpone the TR transition basic to cooperativity in fish Hb.The possible implications of the findings for aquatic respiration are discussed.Abbreviations ATP adenosine triphosphate - DPG 2,3-diphosphoglycerate (glycerate-2,3-bisphosphate) - GTP guanosine triphosphate - IHP inositol hexaphosphate - NTP nucleoside triphosphates In this paperK _T andK _R are defined as theassociation equilibrium constants instead of dissociation constants (as originally defined by Monod et al. 1965) to facilitate comparison with the Adair constants     

云南山楂茎尖的离体培养及其无性系快速繁殖

用云南山楂(Crataegus scabrifolia(Franch.)Rehd.)成年树茎尖和实生芽两种不同发育阶段的材料为外殖体,诱导它们休眠芽萌动,丛生芽条并诱导芽条生根。实验结果如下:1.以成年态的云南山楂侧芽为外植体,培养在附加IAA 0.1—0.5mg/l+6-BA 1-2mg/l的MS培养基上可诱导芽的萌发;将芽继代培养在附加0.5—1mg/l 6-BA的SH或MS培养基上,40天后芽数增殖4—6倍;将芽条截下置于1/2MS培养基上,附加不同浓度的IAA或IBA,可得到50—80％的生根率。2.以实生芽为外殖体,在相同条件下,则20天后芽数增殖便可获4—6倍;98％以上生根。结果表明:云南山楂的幼年态要比成年态易脱分化和再分化。     

Multiple incompatibilities within populations of Culex pipiens L. in southern France

Strains of Culex pipiens derived from natural populations collected in southern France were crossed to determine their ability to give fertile offspring. Uni- and bi-directional incompatibilities occurred between these strains and each of them had its own crossing properties. Compatibility or incompatibility of a cross seemed to be related to the geographic distance separating the parental colonies, but not to their ecological origin (hypogeous or epigeous). Our results showed evidence for the variation in time of crossing properties of a strain.     

Protein kinase C-dependent and -independent pathways of proto-oncogene induction in human astrocytoma cells

We compared the abilities of the muscarinic agonist carbachol, epidermal growth factor (EGF), and phorbol 12-myristate 13-acetate (PMA) to induce proto-oncogene mRNA accumulation and other cellular responses in normal and protein kinase C-deficient 1321-N1 human astrocytoma cells. PMA, carbachol, and EGF all stimulated rapid accumulation of mRNA for the proto-oncogenes c-fos and c-myc in the normal cells; in the protein kinase C-deficient cells, carbachol and EGF, but not PMA, retained this effect, which was not mimicked by the calcium ionophore A23187. Both carbachol and PMA activated protein kinase C in these cells, as evidenced by the stimulated phosphorylation of an acidic Mr 80,000 protein kinase C substrate protein with phosphoamino acid and peptide map identity. This response was mimicked by several other neurotransmitters in these cells, including epinephrine, histamine, oxotremorine, and serotonin, and was abolished in cells made protein kinase C-deficient by preincubation with high concentrations of PMA. Both PMA and carbachol promoted the phosphorylation of the ribosomal protein S6 and activated an S6 protein kinase in the normal but not in the protein kinase C-deficient cells. EGF, in contrast, did not appear to activate protein kinase C, but promoted the phosphorylation of S6 and activation of the S6 kinase in both normal and protein kinase C-deficient cells. We conclude that, in 1321-N1 cells, induction of c-fos and c-myc mRNA can occur through a protein kinase C-dependent pathway and one or more independent pathways, exemplified by the responses to carbachol and EGF in the protein kinase C-deficient cells.     

Purification and characterization of proteolytic fragments of lipocortin I that inhibit phospholipase A2

Human lipocortin I is a 38.5-kDa phospholipase A2 inhibitor that has been produced in Escherichia coli in large quantities by recombinant DNA technology (Wallner, B.P., Mattaliano, R.J., Hession, C., Cate, R. L., Tizard, R., Sinclair, L.K., Foeller, C., Chow, E.P., Browning, J.L., Ramachandran, K.L., and Pepinsky, R.B. (1986) Nature 320, 77-80). To localize the region within the protein responsible for its inhibitory activity, we generated a series of fragments of the recombinant product by limited proteolysis with elastase and characterized their structure by sequencing and peptide mapping. Five active fragments have been analyzed in detail. The smallest is an 18-kDa fragment derived from the amino-terminal half of lipocortin. Three of the larger fragments contain this region. The fifth fragment is missing 83 amino acids from the amino terminus. A region common to all the active fragments (amino acid residues 97-178) is 70% homologous with the corresponding region from a second member of the lipocortin family which recently was cloned (Huang, K-S., Wallner, B.P., Mattaliano, R.J., Tizard, R., Burne, C., Frey, A., Hession, C., McGray, P., Sinclair, L.K., Chow, E.P., Browning, J.L., Ramachandran, K.L., Tang, J., Smart, J.E., and Pepinsky, R.B. (1986) Cell 46, 191-199) and thus presumably is important for activity. In addition to inhibitory fragments, we have isolated a 3-kDa proteolytic fragment from the amino terminus of lipocortin I that contains the known phosphorylation site for protein-tyrosine kinases. Because of sequence homology of the 3-kDa fragment with biologically active synthetic peptides from pp60v-src and middle T antigen, its release by proteases may represent an important part of the activity of lipocortin.     

The properties of batrachotoxin-modified cardiac Na channels, including state-dependent block by tetrodotoxin

Batrachotoxin (BTX) modification and tetrodotoxin (TTX) block of BTX-modified Na channels were studied in single cardiac cells of neonatal rats using the whole-cell patch-clamp recording technique. The properties of BTX-modified Na channels in heart are qualitatively similar to those in nerve. However, quantitative differences do exist between the modified channels of these two tissues. In the heart, the shift of the conductance-voltage curve for the modified channel was less pronounced, the maximal activation rate constant, (tau m)max, of modified channels was considerably slower, and the slow inactivation of the BTX-modified cardiac Na channels was only partially abolished. TTX blocked BTX-modified mammalian cardiac Na channels and the block decreased over the potential range of -80 to -40 mV. The apparent dissociation constant of TTX changed from 0.23 microM at -50 mV to 0.69 microM at 0 mV. No further reduction of block was observed at potentials greater than -40 mV. This is the potential range over which gating from closed to open states occurred. These results were explained by assuming that TTX has a higher affinity for closed BTX-modified channels than for open modified channels. Hence, the TTX-binding rate constants are considered to be state dependent rather than voltage dependent. This differs from the voltage dependence of TTX block reported for BTX-modified Na channels from membrane vesicles incorporated into lipid bilayers and from amphibian node of Ranvier.     

Myasthenogenicity of human acetylcholine receptor synthetic alpha-subunit peptide 125-147 does not require intramolecular disulfide cyclization

This study reports the synthesis of a disulfide-looped peptide corresponding to residues 125-147 (Cys 128-Cys 142) of the nicotinic acetylcholine receptor (AChR) of human skeletal muscle, H alpha 125-147 (Lys-Ser-Tyr-Cys-Glu-Ile-Ile-Val-Thr-His-Phe-Pro-Phe-Asp-Glu-Gln- Asn-Cys-Ser-Nle-Lys Leu-Gly), and a nondisulfide-looped analogue, H alpha 125-147(S) (Lys-Ser-Tyr-Ser-Glu-Ile-Ile-Val-Thr-His-Phe-Pro-Phe-Asp-Glu- Gln-Asn-Cys-Ser-Nle-Lys-Leu-Gly), in which the amino acid Cys 128 was replaced with serine. Both peptides induced antigen-specific helper T cell responses, as evidenced in vitro by lymph node cell proliferation and in vivo by production of anti-AChR antibodies. Rats immunized with 100 micrograms of either synthetic peptide, without conjugation to a carrier, produced anti-peptide antibodies which bound to native AChR in immunoprecipitation assays and induced modulation of membrane-bound AChR from cultured human myotubes. Both peptides also induced electrophysiologic and biochemical signs of experimental autoimmune myasthenia gravis. Thus, region 125-147 of the AChR alpha-subunit is at least partly exposed extracellularly in human muscle and contains one or more autoantigenic sites capable of stimulating T cells and B cells. Disulfide-linkage between residues Cys 128 and Cys 142 is not essential for myasthenogenicity.     

Conservation of δ-crystallin gene structure between ducks and chickens

A cloned chicken delta-crystallin cDNA was used to identify two putative delta-crystallin genes in the duck by Southern blot hybridization. A DNA fragment containing most of one of these genes was isolated from a library made in bacteriophage lambda Charon 28A containing genomic DNA from 14-day-old embryonic ducks. Electron microscopy, partial gene sequencing, primer extension analysis using duck mRNA, and comparison with the well-characterized chicken delta-crystallin genes suggest that our cloned duck delta-crystallin gene, like the chicken delta-crystallin genes, is 8-10 kb long and contains 17 exons. Hybridization and sequencing data show great similarity between the homologous 5' untranslated and coding exons of the duck and chicken delta-crystallin genes. Overall, the homologous introns also appear to have approximately 30% sequence similarity, and have been subject to deletion/insertion events. Our partial characterization of duck delta-crystallin gene sequences suggests that this avian and reptilian crystallin family has been conserved during evolution, as have the other crystallin gene families that are expressed in the eye lens.     

墨红鲜花香气(头香)成分分析

利用 XAD-4憎水性吸附树脂采集墨红头香,以毛细管气相色谱双柱保留指数和 GC/MS/DS 联用方法鉴定头香的化学成份。共分离鉴定或初步鉴定了45种组份,其中含量较大的有乙酸芳樟酯(14.98%),柠檬烯(12.07%),甲基苯甲醚(9.88%),香茅醇(4.82%),乙酸巳酯(3.98%),β-石竹烯(4.55%),芳樟醇(3.18%),正巳醇(3.17%)等.