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Human erythrocytes were treated by a series of SH-reagents, including maleimides, iodo compounds, mercurials and oxidizing agents. Rates of Li efflux into Na-rich medium, Li leak and Lii-Nao countertransport were then determined. Of the 13 different reagents studied, only N-ethylmaleimide, iodoacetamide and iodoacetate inhibited selectively the countertransport activity. The effect of the various reagents indicates that the sensitive SH-groups of the countertransport system are not externally exposed. N-Ethylmaleimide was used to probe for changes elicited by substrate cations in Lii-Nao countertransport. In Na- and Li-free medium, inhibition of Lii-Nao countertransport by N-ethylmaleimide of 35% was reached within 2 s. In Na or Li medium, maximal inhibition was twice as great, but was attained much more slowly, within 10 min. Kinetic data and Hill plot analysis indicate the involvement of two classes of SH-groups: one expressed in the various media with and without substrate cations, and an additional one, which becomes specifically available to N-ethylmaleimide in the presence of external Na or Li. The affinity of Na to the site promoting inhibition by N-ethylmaleimide (apparent Km 12 mM) is higher than the affinity of Na to its external countertransport site (apparent Km 25 mM), as reported by Sarakadi, B., Alifimoff, J.K., Gunn, R.B. and Tosteson, D.C. (1978) J. Gen. Physiol. 72, 249–265). Reactivity of was not modified by the media tested. It is concluded that external Na and Li cause a conformational change in the protein(s) of the countertransport system in human erythrocytes. 相似文献
44.
The structure of the Haemophilus influenzae type f capsular polysaccharide was studied by chemical and nuclear magnetic resonance spectroscopic techniques. The repeating unit of the polysaccharide was found to be . 相似文献
45.
The effect of inflorescence removal on stem elongation in Chinese cabbage cv. Spring A was studied. Removal of the inflorescence before its visibility, or upon its appearance but before the beginning of bolting (stages 1–3), markedly reduced the stem length. Removal after the beginning of bolting (stage 5) had no effect on stem length.Application of GA3 to the treated plants partially or fully restored the elongation of the flowering stem, whereas paclobutrazol inhibited the elongation of the treated, as well as the control stems. Indole-3-acetic acid (IAA) or kinetin was ineffective in restoring stem elongation of the plants from which the inflorescence had been removed. Inflorescences at stages 1–2 were found to secrete about 10 times more gibberellic acid (GA)-like activity compared with control apices or inflorescences at stage 5.It is suggested that the developing inflorescence is the major source of GAs which control stem elongation. However, shortly after the appearance of the inflorescence at the onset of bolting, stem elongation is no longer dependent on GAs derived from the apical inflorescence but require GAs from other sources.Contribution from the Agricultural Research Organization, The Volcani Center Bet Dagan, Israel No. 2218-E, 1987 series. 相似文献
46.
Immunofluorescence microscopy has been used to demonstrate that X22, a monoclonal antibody specific for clathrin heavy chain, localizes in repetitive bands that appear soon after the fusion of skeletal myoblasts into multinucleate fibers. This organization has been found in cultures containing myotubes that develop in vitro from explants of newborn rat hindlimb cells and in myotubes derived from the L8E63 myogenic line. Bands were also prominent in skinned fibers prepared from adult rat soleus muscle and in cardiac myocytes grown in vitro from 4-day heart ventricles. Immunofluorescence banding was localized in the sarcomere as a doublet, with one element on either side of the Z line. Evidence that supports the conclusion that the reaction with X22 antibody is specific and indicative of the localization of clathrin in the sarcomere includes: (1) Identical titration of X22 antibody reactivity with the determinant in coated vesicles and in the sarcomere. (2) Conditions (eg., pH and Tris) that disrupt clathrin baskets or prevent its assembly likewise disrupt the localization of X22 in bands. (3) Chloroquine inhibits both the normal trafficking of clathrin in the cell and X22 banding in the sarcomere. (4) Immunoblot analysis of myotube lysates reveals a single band with an electrophoretic mobility identical to the 180,000-Da clathrin heavy chain. (5) The assembly of clathrin into sarcomeric bands occurs early in the development of the myofibrillar apparatus. Quantitation of the appearance of X22 banding in primary cultures of myotubes indicates that it precedes that of other myofibrillar proteins and that assembly takes place in the following order: X22, titin, myosin heavy chain, actin, and desmin. The assembly of myosin, titin, and actin into sarcomeric bands, as well as X22, is inhibited by chloroquine. Upon prolonged exposure to chloroquine previously assembled proteins are drastically reduced or no longer evident in the sarcomere. On the basis of these results and considering the role of clathrin in intracellular transport and its capacity to interact with actin and alpha-actinin, we suggest that clathrin may have diverse roles in the assembly, integrity, and functioning of the sarcomere and its integration with the sarcolemma. The early organization of X22 into bands further suggests that clathrin may also function early in the assembly of the contractile system. 相似文献
47.
Sulphated polysaccharides and zona pellucida glycoproteins have been shown to bind non-enzymatically to proacrosin, the protein found within the acrosomal vesicle of mammalian spermatozoa. The mechanism of this interaction has been investigated using 125I-fucoidan to probe purified ram sperm proacrosin. Results show that (a) binding of' 125I-fucoidan to proacrosin is inhibited only by sulphated polymers and (b) recognition is mediated by poly(sulphate) groups and is largely independent of the composition of the polymer chain. It is suggested that a similar mechanism is responsible for the interaction between proacrosin and zona pellueida glycoproteins during the early stages of fertilization in mammals and this process mediates firm binding of spermatozoa to the egg. 相似文献
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49.
The proteins of the secretory granules of the rat parotid gland were characterized by sodium dodecylsulfate gel electrophoresis, by chromatography of [3H]prolinelabeled proteins on DEAE-cellulose and by amino acid analysis.Sodium dodecylsulfate gel electrophoresis of the secretory granule content showed five principal proteins and a limited number of minor components. Only two of the principal bands could be identified as known secretory enzymes of the parotid gland. One was identified as the α-amylase and one as deoxyribonuclease. Peroxidase and ribonuclease form minor portions of the secretory proteins.The other three major proteins constitute, together, about 60% by weight, of the secretory granule content proteins. Of these, one which represents more than 30% of the total granule protein was found to contain uniquely high amounts of leucine residues (21 mole%). Another one of these principal proteins was relatively rich in cysteine residues (7 mole%).The fifth principal protein was found to contain high amounts of proline (28 mole%) glutamic acid (17 mole%) and glycine (18 mole%) residues. Its amino acid composition was very similar to that of the proline-rich proteins that were previously shown to be present in the membrane isolated from these granules. This protein, however, differed from the “membranous” proline-rich proteins by several criteria.Two minor glycoproteins of the secretory granule content were also found to be rich in proline residues (37 mole%). As with the other proline-rich proteins of the granule, they contained no sulphur-containing amino acids, stained faintly pink with Coomassie Blue and were underestimated by the Lowry method. They differ however, from all the other proline-rich proteins of the granule by having a significantly higher content of threonine, less glycine (9 mole%) and much less glutamic acid (3 mole%).Of the principal proteins, only the deoxyribonuclease and the half-cystine-rich proteins were positively stained by periodic acid Schiff staining.The possible functions of the leucine-rich, the half cystine-rich and the various proline-rich proteins are discussed. 相似文献
50.
Polyacrylamide gel electrophoresis has been used to analyse basic protein changes during the final stages of spermiogenesis in the house cricket. Mature sperm were obtained from the spermathecae of inseminated females. Their basic protein is electrophoretically heterogeneous, with two major and two minor components, all of unusually high mobilities, as expected ofprotamine. No histones are present. Testis also contains basic protein components of high mobilities, although in small amount relative to the histones present. Testis preparations were centrifuged on a density gradient of colloidal silica to separate nuclei of different stages of spermiogenesis from each other, and it was found that very late spermatids contain relatively large amounts of protamine. At least seven different protamine-like components, each with a different mobility, occur during the final maturation stages. The particular components present, and their abundancies, vary during development. The complement first found in spermatids is different from that of a later spermatid; still another complement is found in sperm from the seminal vesicle; and still another in mature sperm. Components which are abundant in spermatids are progressively eliminated, while components which are barely detectable in them gradually increase in abundance to become the major components of the basic protein complement at maturity. 相似文献