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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Escherichia coli replication factor Y (protein n') functions in the assembly of a mobile multiprotein replication-priming complex called the primosome. Although the role of factor Y in primosome assembly during replication in vitro of bacteriophage phi X174 and plasmid pBR322 DNA is clear, its role in E. coli chromosomal replication is not. To address this issue, the gene for factor Y has been cloned molecularly and its DNA sequence has been determined. The cloned fragment of DNA contained an open reading frame capable of encoding a polypeptide of 81.7 kDa. This open reading frame contains amino acid sequences identical to 13 N-terminal amino acids of purified factor Y, as well as to a 10-amino acid internal sequence (from a cyanogen bromide fragment) as determined by gas-phase microsequencing. Expression of the polypeptide encoded by this open reading frame using a bacteriophage T7 transient expression system resulted in the accumulation of a polypeptide with an apparent molecular mass of 78 kDa that comigrated with bona fide factor Y during SDS/polyacrylamide gel electrophoresis. Soluble extracts made from cells overexpressing the product of the putative factor Y open reading frame showed a 2000-fold increase in factor Y activity during bacteriophage phi X174 complementary-strand DNA synthesis in vitro when compared to control extracts. The gene encoding factor Y, which maps to 88.5 min on the E. coli chromosome, has been designated primosome A (priA).
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PMID:Molecular cloning and DNA sequence analysis of Escherichia coli priA, the gene encoding the primosomal protein replication factor Y. 216 49

A monoclonal antibody, designated mAb P86/5, was generated by immunization of female Balb/c mice with a membrane vesicle fraction composed of the outer acrosomal membrane and plasma membrane (PM-OAM). As determined by fluorescence microscopy and electron microscopy P86/5 recognizes a sperm plasma membrane antigen that is restricted to the sperm head. In intact spermatozoa the P86/5-antigen is distributed over the surface of the sperm head with the exception of the rostral region. By comparing the antibody binding pattern generated at 4 degrees C and 25 degrees C, it could be shown that the P86/5-antigen is capable to diffuse freely within the cell membrane overlying the acrosome whereas its lateral mobility is restricted to the post-acrosomal region. The P86/5-antigen had a molecular weight of about 78 kDa as revealed by SDS-PAGE and western blotting. The glycoprotein nature of the P86/5-antigen was established by lectin affinity chromatography.
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PMID:Boar sperm membranes antigens. I. Topography of a mobile glycoprotein of the sperm cell membrane. 218 99

We examined the qualitative patterns of protein synthesis in fully grown prophase-blocked oocytes of Xenopus laevis and after meiosis reinitiation accompanying maturation of the oocytes. Newly synthesized proteins labelled with [35S]methionine were run on isoelectric focusing gels and further separated in the second dimension on SDS-polyacrylamide slab gels. Three types of maturation inducer were compared: progesterone, considered as the natural inducer of Xenopus oocyte maturation, hCG (human chorionic gonadotropin) and insulin. Three polypeptides with apparent molecular masses of 37 kDa (pI 4.7-4.8), 78 kDa (pI 4.7) and 138 kDa (pI 4.6-4.7) were found to be always synthesized in all three types of stimulation, while the synthesis of a fourth one (molecular mass 116 kDa, pI 4.7) was arrested during oocyte maturation. Moreover, when the follicular cells surrounding the oocytes were part of the stimulating pathway, which is the case during hCG-induced maturation, an additional polypeptide was synthesized by the oocytes (molecular mass 106 kDa, pI 6.0-6.2). This polypeptide was not synthesized during progesterone- or insulin-induced oocyte maturation, two types of stimulation which do not require the presence of the follicular cells. The biological significance of the hCG-induced polypeptide, not necessary for oocyte maturation, is discussed. On the other hand, the four other modifications in protein synthesis taking place during all three types of maturation-inducing stimulation appear to be necessary for oocyte maturation, since oocytes which failed to mature in response to stimulation always missed one or several of these four polypeptides.
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PMID:Patterns of protein synthesis during Xenopus oocyte maturation differ according to the type of stimulation. 227 96

Neuroblastoma x glioma hybrid cells (NG108-15) were used as a model system to characterize neuronal-glial type angiotensin (ANG) receptors by covalent crosslinking analysis. After differentiation with 1.5% DMSO and 0.5% fetal bovine serum for four to five days, saturation analysis revealed a single high affinity site with a Kd = 1.35 +/- 0.42 nM and a Bmax = 468 +/- 106 fmol/mg protein. Using the homobifunctional crosslinking reagent bis(sulfosuccinimidyl) suberate (BS3), a site with an estimated Mr of 78 kDa was specifically labeled with 125I-ANG II as determined by SDS-polyacrylamide gel electrophoresis. Both ANG II and ANG III (10(-6) M) inhibited specific labeling. The Ki for ANG III binding was similar by both pharmacologic (Ki = 3.33 +/- 0.98 nM) and gel densitometric (Ki = 2.65 +/- 0.32 nM) analyses. We conclude that the 78 kDa protein represents a high affinity ANG binding site with similar affinities for both ANG II and ANG III.
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PMID:Covalent crosslinking analysis of angiotensin receptors on differentiated NG108-15 cells. 239 77

Proteins of the C1 complex, C1q, C1r, and C1s, of the classical pathway of complement activation are known to be synthesized in human skin fibroblasts. Using metabolic labeling with [35S]methionine, immunoprecipitation, and SDS-PAGE, we demonstrate that human skin fibroblasts synthesize and secrete C1 inhibitor with an apparent molecular mass of 78 kDa in the cell lysate and 102 kDa in the extracellular medium. This C1 inhibitor had the capacity to bind activated C1s. Fibroblasts synthesized 30- to 50-fold more C1 inhibitor than was synthesized in monocytes. As previously reported, fibroblasts also synthesized C1r and C1s. IFN-gamma, IFN-beta 1, and TNF had significant, but distinct, effects on synthesis of C1 inhibitor, C1r, and C1s. Incubation of the cells with IFN-gamma, 1000 U/ml, for 24 h induced increases in the synthesis of C1 inhibitor, C1r, and C1s by 4.2-, 1.9- and 1.6-fold, respectively. IFN-beta 1 had effects similar to IFN-gamma, although smaller in magnitude. TNF, 12.5 ng/ml, induced increases in the synthesis of C1 inhibitor, C1r, and C1s by 1.5-, 1.4- and 2.6-fold. IL-1, IFN-beta 2 (IL-6), and LPS did not affect synthesis of C1 inhibitor, C1r, or C1s. Fibroblasts are present in large amounts in most tissues. Synthesis of C1 inhibitor, C1r, and C1s by these cells could provide a source of these important proteins in body tissues. In addition, fibroblasts should be a good model for the in vitro study of genetic diseases involving the synthesis of these proteins.
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PMID:Synthesis and regulation of C1 inhibitor in human skin fibroblasts. 253 70

We have determined that the major iron-binding and DNA-binding protein in porcine colostral whey is lactoferrin. This lactoferrin was purified to homogeneity in one chromatographic step using immobilized single-stranded DNA-agarose. Although different in chromatographic behavior from human lactoferrin, the porcine lactoferrin purified in this manner was shown to be homogeneous by high-performance ion-exchange chromatography (Mono-S), immobilized metal ion (Cu2+) affinity chromatography, size-exclusion chromatography (TSK-4000SW), and reverse-phase (phenyl) chromatography. Electrophoresis on SDS-polyacrylamide gradient (10-20%) gels under reducing conditions showed the purified lactoferrin to be a single protein (silver-stained) of 78 kDa. Apolactoferrin purified in this manner bound iron and displayed a UV/VIS absorption spectrum indistinguishable from that of human lactoferrin. The molar absorption coefficient of hololactoferrin was 3.86 x 10(3) M-1 at 465 nm and 1.08 x 10(5) M-1 at 280 nm. Affinity elution analyses of the purified lactoferrin on immobilized DNA revealed that the affinity of this protein for DNA was independent of bound iron. Porcine lactoferrin was recognized by antibodies directed against human lactoferrin and bovine lactoferrin. The amino acid composition and N-terminal amino acid sequence analysis (30 residues) revealed a high degree of sequence homology with human, equine and bovine lactoferrin. These results demonstrate the effectiveness of immobilized DNA as a rapid and simple lactoferrin purification procedure and demonstrate the presence of a lactoferrin in porcine colostral whey with a high degree of sequence homology to human lactoferrin.
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PMID:Rapid purification of porcine colostral whey lactoferrin by affinity chromatography on single-stranded DNA-agarose. Characterization, amino acid composition and N-terminal amino acid sequence. 260 66

Little is known about the metabolic fate of ingested lactoferrin in human-milk-fed term or preterm infants. Enzyme-linked immunosorbent assays and sodium dodecyl sulfate/polyacrylamide gradient gel electrophoresis (SDS-PAGGE) with immunoblots have demonstrated that the urinary excretion of lactoferrin by preterm infants fed exclusively human milk exceeded that by formula-fed infants. The origin and molecular integrity of the excreted lactoferrin, however, are unclear. We have developed extraction and separation procedures involving a stationary phase of immobilized single-stranded DNA (ssDNA) that allows the efficient (greater than 80%) and rapid isolation of pure, intact lactoferrin from infants' urine. We purified lactoferrin to apparent homogeneity from infants' urine in one step, using the immobilized ssDNA with mobile phases containing up to 6 mol of urea per liter. The purified lactoferrin was evaluated by SDS-PAGGE; silver-staining revealed one protein band at 78 kDa; immunoblots confirmed the presence of intact lactoferrin. High-performance affinity chromatography with use of immobilized metal ion (Cu2+) suggested the presence of intact, iron-saturated lactoferrin. Subsequent chromatography on high-performance reversed-phase (C18) columns independently verified sample identity and purity.
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PMID:Purification and characterization of intact lactoferrin found in the urine of human milk-fed preterm infants. 277 18

Thrombin (Th) binds specifically to confluent cultures of adult bovine aortic (ABAE) and bovine brain capillary (BBC) endothelial cells. Saturation of 125I-Th binding is observed after 1 h exposure to the ligand and at an extracellular concentration of 0.5 and 1.0 microgram/ml for ABAE and BBC cells, respectively. Under optimal conditions both ABAE and BBC cultures bind about 2 to 5 ng/10(6) cells, which represents about 20% of Th binding to bovine corneal endothelial (BCE) cells. Under optimal conditions less than 30% of the total cell associated 125I-Th is internalized in ABAE and BBC cells, while in BCE cells the extent of internalization is more than 50%. The internalized 125I-Th is degraded both in ABAE and BBC cells as previously demonstrated in BCE cells. As analyzed by SDS-PAGE, 17%, 22% and 77% of the bound 125I-Th is in complex with anti-thrombins (anti-Ths) in BBC, ABAE and BCE cultures, respectively. ABAE cells possess 3 types of complexes, one which appears only on the cell surface with a molecular weight of 78 kDa, and two others which appear only in the conditioned medium (CM) with molecular weights of 84 and 85 kDa. BBC and BCE cells demonstrate only one type of complex with a molecular weight of 77 kDa which appears both on the cell surface and in the CM. The 125I-Th 77 kDa complex formed in the CM of BCE cells is recognized and bound by BBC cells and ABAE cells.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Anti-thrombin activity in cultured aortic and capillary endothelial cells: binding, internalization and degradation of thrombin. 277 78

Because of its anatomy, the neuron R2 of Aplysia has been used to study how proteins are distributed to their appropriate destinations within the cell. The R2 cell body resides in the abdominal ganglion, while its axons terminate on glands in the skin. Using intracellular injection of HRP and intraxonal recordings, we found that R2 has a dendritic (receptive) arborization in the pleural ganglion. The structure of these dendrites was examined after injecting the soma with 3H-L-fucose, thereby labeling glycoproteins that are transported to all regions of the cell. Light- and electron-microscope autoradiography show that the openings to the dendrites are not on the periphery, but are suspended inside the axon by glial cell infoldings. All of the organelles seen in the axon are found in the dendrites, including 2 types of vesicles. Neither the axon nor the dendrites contain ribosomes. Thus, R2 has 3 functionally distinct regions--cell body, dendrites, presynaptic terminals--that are separated from each other by at least 4 cm. This implies that pre- and postsynaptic proteins made in the cell body are transported along the axon to the pleural ganglion, where they are sorted. To investigate this idea, we exposed the abdominal ganglion to 35S-methionine to label R2's proteins. Analyses by SDS-PAGE of the rapidly transported labeled proteins from R2 consistently showed a 78 kDa band that accumulated in the pleural ganglion and did not move into the peripheral nerves. This then is a putative dendritic constituent.
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PMID:Giant Aplysia neuron R2 has distal dendrites: evidence for protein sorting and a second spike initiation site. 282 69

A heparin-binding protein was isolated from bovine uteri and purified to homogeneity. This protein appears as a double band of approx. 78 kDa in SDS/polyacrylamide-gel electrophoresis and has an isoelectric point of 5.2. The binding of heparin to this protein is saturable. No other glycosaminoglycan from mammalian tissue, such as hyaluronic acid, chondroitin sulphate, dermatan sulphate or keratan sulphate, binds to the 78 kDa protein. Dextran sulphate binds in a non-saturable fashion. Certain heparan sulphate polysaccharide structures are required for binding to the 78 kDa protein. Some proteoheparan sulphates, such as endothelial cell-surface proteoheparan sulphate, show only weak interaction with the 78 kDa protein in contrast with a basement-membrane proteoheparan sulphate from HR-9 cells. Antibodies against the 78 kDa protein inhibit binding of proteoheparan [35S]sulphate from basement membranes to smooth-muscle cells. Conventional antibodies, Fab fragments and some monoclonal antibodies, inhibit smooth-muscle cell proliferation in a similar range as that observed for heparin. The protein was detected in a variety of tissues and cells but not in blood cells. A possible role of this protein as a receptor for heparin or heparan sulphate and its function in the control of the arterial wall structure are discussed.
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PMID:A heparin-binding protein involved in inhibition of smooth-muscle cell proliferation. 304 3


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