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61.
我们分子鉴别了一个缺失型中国(A_γδβ)°-地贫家系。先证者为这一缺失的纯合子,具有中度贫血症状。家系的另五个成员均为这一缺失的杂合子,其胎儿血红蛋白(HbF)为16—21%,接近或达到HPFH杂合子的HbF水平,并且几乎不表现贫血症状。限制性内切酶图谱分析证明了β-珠蛋白基因簇内的DNA顺序缺失,缺失的5′端点位于Aγ基因IVSⅡ内,3′端点在β-珠蛋白基因下游区远端,距HPFH-2的3′缺失端点上游区约11kb。缺失的总长度约为80kb。本文讨论了这一缺失导致胎儿血红蛋白在成人中持续活跃表达的可能机制。 相似文献
62.
Studies with a rolling-circle DNA replication system reconstituted in vitro with a tailed form II DNA template, the DNA polymerase III holoenzyme (Pol III HE), the Escherichia coli single-stranded DNA binding protein, and the primosome, showed that within the context of a replication fork, the oligoribonucleotide primers that were formed were limited to a length in the range of 9 to 14 nucleotides, regardless of whether they were subsequently elongated by the lagging-strand DNA polymerase. This is in contrast to the 8-60-nucleotide-long primers synthesized by the primosome in the absence of DNA replication on a bacteriophage phi X174 DNA template, although when primer synthesis and DNA replication were catalyzed concurrently in this system, the extent of RNA polymerization decreased. As described in this report, we therefore examined the effect of the DNA Pol III HE on the length of primers synthesized by primase in vitro in the absence of DNA replication. When primer synthesis was catalyzed either: i) by the primosome on a phi X174 DNA template, ii) by primase on naked DNA with the aid of the DnaB protein (general priming), or iii) by primase alone at the bacteriophage G4 origin, the presence of the DNA Pol III HE in the reaction mixtures resulted in a universal reduction in the length of the heterogeneous RNA products to a uniform size of approximately 10 nucleotides. dNTPs were not required, and the addition of dGMP, an inhibitor of the 3'----5' exonuclease of the DNA Pol III HE, did not alter the effect; therefore, neither the 5'----3' DNA polymerase activity nor the 3'----5' exonuclease activity of the DNA Pol III HE was involved. E. coli DNA polymerase I, and the DNA polymerases of bacteriophages T4 and T7 could not substitute for the DNA Pol III HE. The Pol III core plays a crucial role in mediating this effect, although other subunits of the DNA Pol III HE are also required. These observations suggest that the association of primase with the DNA Pol III HE during primer synthesis regulates its catalytic activity and that this regulatory interaction occurs independently of, and prior to, formation of a preinitiation complex of the DNA Pol III HE on the primer terminus. 相似文献
63.
The carbohydrate specificity of mistletoe toxic lectin-I (ML-I) was studied by haemagglutination-inhibition assay. The results indicated that ML-I has a broad range of affinity for Gal alpha,beta linked sequences. The galabiose (E, Gal alpha 1----4Gal) sequence, a receptor of the uropathogenic E. coli ligand, was one of the best disaccharide inhibitors tested. The lectin also exhibits affinity for Lac(Gal beta 1----4Glc), T(Gal beta 1----3GalNAc), I/II(Gal beta 1----3/4GlcNAc) and B(Gal alpha 1----3Gal) sequences. Gal alpha 1----4Gal and Gal beta 1----4Glc are frequently occurring sequences of many glycosphingolipids located at the mammalian cell membranes, such as intestinal and red blood cell membranes, for ligand binding and toxin attachment. This finding provides important information concerning the possible mechanism of intoxication of cells by the mistletoe preparation. 相似文献
64.
K Ohashi K H Ruan R J Kulmacz K K Wu L H Wang 《The Journal of biological chemistry》1992,267(2):789-793
Polymerase chain reaction techniques have been used to isolate a cDNA clone containing the entire protein coding region of thromboxane A2 synthase (EC 5.3.99.5) from a human lung cDNA library. The cDNA clone hybridizes with a single 2.1-kilobase mRNA species in phorbol ester-induced human erythroleukemia and monocytic leukemia cell lines. A second cDNA, differing only by an insert of 163 base pairs near the 3'-end of the translated region, was also found to be present in the same library. The proteins predicted from both nucleic acid sequences include the three polypeptide sequences determined from amino acid sequencing of the purified human platelet enzyme, five potential sites for N-glycosylation, and a hydrophobic region that may serve to anchor the synthase in the endoplasmic reticulum membrane. The longer predicted protein, designated thromboxane synthase-I, contains 534 amino acids, with a Mr of 60,684, whereas the shorter protein, designated thromboxane synthase-II, contains 460 amino acids and has a Mr of 52,408. Although thromboxane synthase-II lacks the conserved cysteine that serves as the proximal heme ligand in the other cytochromes, significant sequence similarities exist among thromboxane synthase-I and -II and several P450s, particularly those in family 3. The overall amino acid identity is considerably less than 40%, making it likely that thromboxane synthase represents a previously undefined family of cytochrome P450. 相似文献
65.
T-cell receptor (Tcr) chains are classified into four subgroups (I, II, III, and miscellaneous) based on the amino acid residues at positions 61 and 62. Subgroup I has Gly Phe at these positions, subgroup II has Arg Phe, subgroup III has Arg Leu, and subgroup miscellaneous has several other combinations. Variability plots for subgroups I, II, and III sequences show higher values around positions 93–103, 105, 108, 111, 113, and 115, suggesting that these positions may interact with the processed antigen molecules. Smaller peaks are present at various other regions which may bind the major histocompatibility complex class I or II molecules. The patterns of variability within one subgroup are similar for all species, for human alone, and for mouse alone. These subgroup patterns appear much less complicated than patterns for sequences in all subgroups taken together, implying that subgroups may be related to Tcr functions. Among 83 mouse chains, 15 are from cytotoxic cells and 40 from helper cells. Of the 15 from cytotoxic cells, 11, 2, 0, and 2 are in subgroups I, II, III, and miscellaneous; and of the 40 from helper cells, 9, 16, 12, ans 3 are in subgroups I, II, III, and miscellaneous, respectively. Thus, a correlation between sequence and function of Tcr chains seems possible.
Address correspondence and offprint requests to: M. Schiffer. 相似文献
66.
67.
68.
S T Butera V L Perez N J Besansky W C Chan B Y Wu G J Nabel T M Folks 《Journal of cellular biochemistry》1991,45(4):366-373
In this report, we describe a human immunodeficiency virus type-1 (HIV-1)-infected promyelocytic cell line, OM, derived from HL-60 cells. Although the OM cell line was biologically cloned twice, the pattern of HIV-1 expression during culture appeared analogous to a classical acute spreading infection and was inhibited by both azidothymidine and recombinant soluble CD4 treatment. The number of OM cells actually expressing HIV-1 at the beginning of culture was 0%, reached a peak of nearly 100% at 6 weeks, and then fell to less than 10% HIV-1+ cells by 10 weeks. Clonal analysis of the surviving cells verified that stable HIV-1+ OM cells resulted from the spreading infection. Southern analysis confirmed the transmission of HIV-1 through these OM cultures and the occurrence of stable clones which resulted. The initial percentage of OM cells actually harboring the HIV-1 genome was less than 0.1%, indicating nonfaithful transmission of an unintegrated HIV-1 genome during clonal expansion. These results demonstrate that extrachromosomal HIV-1 DNA can contribute to the spread of HIV-1 infection and give rise to cells which have stably integrated HIV-1 provirus. 相似文献
69.
Summary An Enzyme Linked Immuno Sorbent Assays (ELISA) and an Electro-chemiluminescent Immuno Assay (CIA) are compared for the purpose of monitoring product formation in mammalian cell bioreactors. The ELISA had a relative standard deviation of 10%, compared to 8% for the CIA . The CIA was found to be a fast and accurate alternative to the ELISA. The use of more than one immuno assay format was also shown to provide additional insight into process performance. 相似文献
70.
We present a new method for determining the orientation of alpha-helical sections of proteins or peptides in membrane. To apply this method, membranes containing proteins must be prepared in a multilayer array. Circular dichroism (CD) spectra of the multilayer sample are then measured at the normal as well as oblique incident angles with respect to the bilayer planes; we call such spectra oriented circular dichroism (OCD). The procedure of OCD measurement, particularly the ways to avoid the spectral artifacts due to the effects of dielectric interfaces, linear dichroism and birefringence, and the method of data analysis are described in detail. To illustrate the method, we analyze the OCD of alamethicin in diphytanoylphosphatidylcholine multilayers. We conclude unambiguously that the helical section of alamethicin is parallel to the membrane normal when the sample is in the full-hydration state, but the helical section rotates to the plane of membrane when the sample is in a low-hydration state. We also obtained the parallel and perpendicular CD spectra of alpha-helix, and found them to be in agreement with previous theoretical calculations based on the exciton theory. These spectra are useful for analyzing protein orientations in future experiments. 相似文献