共查询到20条相似文献,搜索用时 15 毫秒
1.
Pancreatic hormones have been purified from the opossum, a New World marsupial. Opossum insulin contains a Leu substitution at the N-terminus of the B-chain in place of the Phe that is generally present in mammalian insulins. In addition, there are two other amino acid substitutions in the opossum insulin A-chain (positions 8 and 18) compared to pig insulin. Opossum glucagon is identical to chicken glucagon with both differing from the usual mammalian glucagon by Ser in place of Asn at its penultimate C-terminal position. Opossum PP differs from the porcine peptide in only 3 sites (position 3, 19 and 30). 相似文献
2.
Angela Chambery Antimo Di Maro Maurilia M Monti Fiorenzo Stirpe Augusto Parente 《European journal of biochemistry》2004,271(1):108-117
Volkensin, a type 2 ribosome-inactivating protein from the roots of Adenia volkensii Harms (kilyambiti plant) was characterized both at the protein and nucleotide level by direct amino acid sequencing and cloning of the gene encoding the protein. Gene sequence analysis revealed that volkensin is encoded by a 1569-bp ORF (523 amino acid residues) without introns, with an internal linker sequence of 45 bp. Differences in residues present at several sequence positions (reproduced after repeated protein sequence analyses), with respect to the gene sequence, suggest several isoforms for the volkensin A-chain. Based on the crystallographic coordinates of ricin, which shares a high sequence identity with volkensin, a molecular model of volkensin was obtained. The 3D model suggests that the amino acid residues of the active site of the ricin A-chain are conserved at identical spatial positions, including Ser203, a novel amino acid residue found to be conserved in all known ribosome-inactivating proteins. The sugar binding site 1 of the ricin B-chain is also conserved in the volkensin B-chain, whilst in binding site 2, His246 replaces Tyr248. Native volkensin contains two free cysteinyl residues out of 14 derived from the gene sequence, thus suggesting a further disulphide bridge in the B chain, in addition to the inter- and intrachain disulphide bond pattern common to other type 2 ribosome-inactivating proteins. 相似文献
3.
Identification of a cell retention signal in the B-chain of platelet-derived growth factor and in the long splice version of the A-chain. 总被引:9,自引:0,他引:9
下载免费PDF全文
![点击此处可从《Molecular biology of the cell》网站下载免费的PDF全文](/ch/ext_images/free.gif)
A Ostman M Andersson C Betsholtz B Westermark C H Heldin 《Molecular biology of the cell》1991,2(7):503-512
The B-chain homodimer of platelet-derived growth factor (PDGF) is only very inefficiently secreted and remains largely associated with the producer cell; in contrast, the dimer of the short, and most common, splice variant of the A-chain is secreted. To identify the structural background to the differences in the secretory pattern between the different isoforms of PDGF, a set of chimeric PDGF A/B cDNAs was generated and expressed in COS cells. Analyses of the biosynthesis and processing of the corresponding products led to the identification of a determinant for cell association in the carboxy-terminal third of the PDGF B-chain precursor. Introduction of stop codons at various positions in the carboxy-terminal prosequence of the PDGF B-chain localized this determinant to an 11-amino-acid-long region (amino acids 219-229). This region contains an 8-amino-acid-long basic sequence that is homologous to a sequence present in an alternatively spliced longer version of the PDGF A-chain. In contrast to the short splice variant, the long splice A-chain version, like the B-chain, was found to remain predominantly cell associated. Thus, we have identified a conserved sequence that inhibits the secretion of some of the PDGF isoforms. Our data also suggest that switching of splicing patterns can be a mechanism to regulate the formation of secreted or cell-associated forms of PDGF-AA and possibly other growth factors. 相似文献
4.
S Stoev S Zakhariev E Golovinsky H G Gattner V K Naithani A Wollmer D Brandenburg 《Biological chemistry Hoppe-Seyler》1988,369(12):1307-1315
The synthesis of [Phe(F)A19]insulin (porcine) is described. First the protected [Phe(F)19]A-chain was assembled by segment condensation of [1-12] and [13-21] using the dicyclohexyldiimide/1-hydroxybenzotriazole procedure. [Phe(F)19]A-chain was purified by ion exchange chromatography after removal of all the protecting groups (Boc, But, OBut and S-Trt) and its conversion into the tetra-S-sulfonated derivative. [Phe(F)A19]insulin was prepared by combination with porcine B-chain and purified by gel filtration and ion-exchange chromatography. The in vitro biological activity of this analogue was 60%. CD spectra in the near and far UV are qualitatively very similar to those of insulin. 相似文献
5.
R Knorr W Danho E E Büllesbach H G Gattner H Zahn G L King C R Kahn 《Hoppe-Seyler's Zeitschrift für physiologische Chemie》1982,363(12):1449-1460
An analogue of porcine insulin which differs from the native molecule in that the amino-acid residue B22-L-arginine is replaced by its D-enantiomer has been synthesized. The [D ArgB22]B-chain was synthesized by the segment condensation method and purified as the di-S-sulfonate by ion exchange chromatoggraphy on SP-Sephadex at pH 3.5. Combination with native porcine sulfhydryl A-chain gave [DArgB22]insulin which was purified by ion exchange chromatography on SP-Sephadex at pH 4.5 with a linear NaCl gradient. The biological activity of this analogue as measured by glucose oxidation in rat epididymal adipocytes was 2%. Thymidine incorporation into DNA of human fibroblast was 16%. The immunoreactivity using antipork insulin antibody in a double antibody immunoassay was 4%. The receptor-binding affinity as measured by radioreceptor assays was 2% with cultured human fibroblasts and 1% with rat adipocytes. These results suggest that the L-configuration at B22-arginine is essential for retaining the biological, immunological and receptor-binding properties of the hormone. 相似文献
6.
Mei HC Li YF Hsu CC Tsai YC Takagi H 《Bioscience, biotechnology, and biochemistry》2003,67(7):1601-1604
Replacement of Gly124 on the S1 pocket of subtilisin YaB with Ala changed the cleavage pattern on oxidized insulin B-chain from the subtilisin type to the elastase type. The initial cleavage site in the B-chain shifted from L15-Y16 for wild-type YaB to A14-L15 for the G124A mutant. Upon complete hydrolysis with the G124A mutant, four of the six major cleavage sites on the B-chain were identical to porcine pancreatic elastase cleavage sites. 相似文献
7.
Isolation and structural characterization of porcine coupling factor 6 from intestinal tissues 总被引:1,自引:0,他引:1
A polypeptide purified from an extract of thermostable, porcine intestinal peptides was found to correspond to coupling factor 6, previously known as a component of the mitochondrial oxidative phosphorylation system. The intestinal presence of this peptide offers a new source for preparation of the component in large quantities, and possibly suggests further functions of the polypeptide. Amino acid sequence analysis of this porcine form reveals it to be identical to the bovine form, except for two replacements, at position 62 (Thr in the porcine, Phe/Thr in the bovine form), and position 70 (Ala/Val). The extensive conservation suggests strict structural constraints on the functional properties of the polypeptide. 相似文献
8.
H J Wieneke W Danho E E Büllesbach H G Gattner H Zahn 《Hoppe-Seyler's Zeitschrift für physiologische Chemie》1983,364(5):537-550
The chemical synthesis of [Tyr(I)A19] and [Tyr(I2)A19]insulin (porcine), using the amino-acid derivatives 3-iodotyrosine and 3,5-diiodotyrosine is described. The synthesis of the iodinated A-chains were performed by segment condensation in solution using acid labile protecting groups. The hydroxyl groups of Tyr(I) and Tyr(I2) were unprotected. For the temporary protection of the alpha-amino groups of the A-chain segments containing iodinated tyrosines, the 1-(4-biphenylyl)-1-methylethoxycarbonyl group was selected. After deprotection and sulphitolysis the iodinated A-chain tetra-S-sulphonates were purified by ion exchange chromatography on DEAE cellulose at pH 5.6. Reduction to the sulphhydryl form and the combination with native porcine B-chain yielded [Tyr(I)A19] and [Tyr(I2)A19]insulin (porcine), respectively. Purification of the first product was achieved by gel filtration and of the later by ion exchange chromatography on CM-cellulose at pH 4.5 and gel filtration. The monoiodinated insulin had a biological activity of 24 +/- 2% and the diiodinated analogue 2.6 +/- 0.2% as determined in an in vitro lipogenesis assay with epididymal adipocytes. 相似文献
9.
本文报道了用Fmoc固相法合成3种胰岛素A链小环(A8-10)被不同碱性氨基酸取代的A链类似物,并分别与天然胰岛素B链重组成相应胰岛素类似物;经受体结合,整体活性及抗体结合实验,均表现出相应的活性。从中可以推测出:A链小环区域不是胰岛素表现生物活性的重要部位,而是胰岛素与其抗体结合较重要的区域。 相似文献
10.
L Xie B Z Wang R G Hu H B Ji L Zhang W Y Liu 《European journal of biochemistry》2001,268(22):5723-5733
Cinnamomin is a new type II ribosome-inactivating protein (RIP). Its A-chain exhibits RNA N-glycosidase activity to inactivate the ribosome and thus inhibit protein synthesis, whereas the glycosylated B-chain is a lectin. The primary structure of cinnamomin, which exhibits approximately 55% identity with those of ricin and abrin, was deduced from the nucleotide sequences of cDNAs of cinnamomin A- and B-chains. It is composed of a total of 549 amino-acid residues: 271 residues in the A-chain, a 14-residue linker and 264 residues in the B-chain. To explore its biological function, the cinnamomin A-chain was expressed in Escherichia coli with a yield of 100 mg per L of culture, and purified through two-step column chromatography. After renaturation, the recovery of the enzyme activity of the expressed A-chain was 80% of that of native A-chain. Based on the modeling of the three-dimensional structure of the A-chain, the functional roles of five amino acids and the only cysteine residues were investigated by site-directed mutagenesis or chemical modification. The conserved single mutation of the five amino-acid residues led to 8-50-fold losses of enzymatic activity, suggesting that these residues were crucial for maintaining the RNA N-glycosidase activity of the A-chain. Most interestingly, the strong electric charge introduced at the position of the single cysteine in A-chain seemed to play a role in enzyme/substrate binding. 相似文献
11.
The position of the disulfide bonds in human plasma alpha 2 HS-glycoprotein and the repeating double disulfide bonds in the domain structure 总被引:1,自引:0,他引:1
The positions of the inter- and intra-chain disulfide bonds of human plasma alpha 2 HS-glycoprotein were determined. alpha 2 HS-glycoprotein was digested with acid proteinase and then with thermolysin. The disulfide bonds containing peptides were separated by reversed-phase HPLC and detected by SBD-F (7-fluorobenzo-2-oxa-1,3-diasole-4-sulfonic acid ammonium salt) method. One inter-disulfide bond containing peptide and five intra-disulfide bond containing peptides (A-chain) were purified and identified as Cys-18 (B-chain)--Cys-14 (A-chain), Cys-71--Cys-82, Cys-96--Cys-114, Cys-128--Cys-131, Cys-190--Cys-201 and Cys-212--Cys-229, respectively. The location of the intra-disulfide bonds revealed that the A-chain of alpha 2 HS-glycoprotein is composed of three domains. Two domains were shown to possess intramolecular homology judging from the total chain length of the domains, size of the loops formed by the S--S bonds, the location of two disulfide loops near the C-terminal end of domains A and B, the distance between two S--S bonds of each domain, the amino acid sequence homology between these two domains (22.6%), number of amino acid residues between the second S--S loops and the end of domains A and B, and the positions of the ordered structures. 相似文献
12.
The cytotoxicity of intact cinnamomin (a type II ribosome-inactivating protein, RIP) and the RNA N-glycosidase activity of cinnamomin A-chain have been studied and compared with those of ricin. Cinnamomin A-chain exhibits a similar RNA N-glycosidase activity in inhibiting in vitro protein synthesis compared with that of ricin, whereas the cytotoxicity to BA/F3beta cells of intact cinnamomin is markedly lower than intact ricin. In order to demonstrate that it is the B-chains of the two RIPs that bear the difference in cytotoxicity, two hybrid RIPs are prepared from the purified A-/B-chains of cinnamomin and ricin by the disulfide exchange reaction. It has been found that hybrid RIP constructed from cinnamomin A-chain and ricin B-chain is more toxic to BA/F3beta cells than the native cinnamomin, and equivalent to the native ricin. However, the cytotoxicity to BA/F3beta cells of the hybrid RIP constructed from the ricin A-chain and cinnamomin B-chain is lower than ricin, equivalent to the native cinnamomin. Furthermore, the bound amounts of two B-chains on the cell surface are determined by the method of direct cellular ELISA and Scatchard analysis of the binding of the two B-chains indicates that cinnamomin and ricin share similar binding sites with different affinity. 相似文献
13.
We studied the mechanism responsible for nonlinear double reciprocal plots for tissue type plasminogen activator (tPA)-mediated plasminogen activation reported previously by several groups. We found nonlinear Eadie-Scatchard plots for Glu-plasminogen activation by recombinant single-chain tPA confirming a non-Michaelis-Menten behavior of tPA. In order to characterize this mechanism, enzyme kinetic studies with truncated substrates (Lys- and miniplasminogen) and modified or truncated enzymes (two-chain tPA and tPA B-chain) were performed. Thereby it could be excluded that product-mediated modifications of the enzyme or the substrate are responsible for the nonlinear plots. Linear plots, i.e., Michaelis-Menten kinetics, were only found when tPA B-chain was used as a plasminogen activator, indicating that the tPA A-chain should be responsible for the non-Michaelis-Menten behavior. Binding studies of plasminogen to immobilized tPA A-chain in fact demonstrated a saturable binding of Glu- and miniplasminogen to the A-chain of tPA with a KD approximately 0.1 microM and one binding site per molecule of tPA A-chain. These data suggested a modifier mechanism responsible for the nonlinear plots whereby the substrate plasminogen itself could function as a modifier. When such a mechanism was included into a model for tPA-mediated plasminogen activation, the experimentally obtained data could be fitted into such a model by nonlinear regression analysis with resulting p-values of less than 0.001. 相似文献
14.
15.
Identification of INSL6, a new member of the insulin family that is expressed in the testis of the human and rat 总被引:6,自引:0,他引:6
Lok S Johnston DS Conklin D Lofton-Day CE Adams RL Jelmberg AC Whitmore TE Schrader S Griswold MD Jaspers SR 《Biology of reproduction》2000,62(6):1593-1599
A new member of the insulin gene family (INSL6) was identified from an Expressed Sequence Tag database through a search for proteins containing the insulin family B-chain cysteine motif. Human and rat INSL6 encoded polypeptides of 213 and 188 amino acids, respectively. These orthologous sequences contained the B-chain, C-peptide, and A-chain motif found in other members of the insulin family. Human INSL6 was 43% identical to human relaxin H2 in the B- and A-chain regions. As with other family members, human and rat INSL6 had predicted dibasic sequences at the junction of the C-peptide and A-chain. Human INSL6 sequence had an additional dibasic site near the C-terminus of the A-chain. The presence of a single basic residue at the predicted junction of the B-chain and C-peptide suggests that multiple prohormone convertases are required to produce the fully mature hormone. INSL6 was found to be expressed at high levels in the testis as determined by Northern blot analysis and specifically within the seminiferous tubules in spermatocytes and round spermatids as detected by in situ hybridization analysis. Radiation hybrid mapping placed the human INSL6 locus at chromosome 9p24 near the placenta insulin-like homologue INSL4 and the autosomal testis-determining factor (TDFA) locus. 相似文献
16.
A common polymorphism in the SFTPD gene influences assembly, function, and concentration of surfactant protein D 总被引:6,自引:0,他引:6
Leth-Larsen R Garred P Jensenius H Meschi J Hartshorn K Madsen J Tornoe I Madsen HO Sørensen G Crouch E Holmskov U 《Journal of immunology (Baltimore, Md. : 1950)》2005,174(3):1532-1538
Surfactant protein D (SP-D) plays important roles in the host defense against infectious microorganisms and in regulating the innate immune response to a variety of pathogen-associated molecular pattern. SP-D is mainly expressed by type II cells of the lung, but SP-D is generally found on epithelial surfaces and in serum. Genotyping for three single-nucleotide variations altering amino acids in the mature protein in codon 11 (Met(11)Thr), 160 (Ala(160)Thr), and 270 (Ser(270)Thr) of the SP-D gene was performed and related to the SP-D levels in serum. Individuals with the Thr/Thr(11)-encoding genotype had significantly lower SP-D serum levels than individuals with the Met/Met(11) genotype. Gel filtration chromatography revealed two distinct m.w. peaks with SP-D immunoreactivity in serum from Met/Met(11)-encoding genotypes. In contrast, Thr/Thr(11) genotypes lacked the highest m.w. form. A similar SP-D size distribution was found for recombinant Met(11) and Thr(11) expressed in human embryonic kidney cells. Atomic force microscopy of purified SP-D showed that components eluting in the position of the high m.w. peak consist of multimers, dodecamers, and monomers of subunits, whereas the second peak exclusively contains monomers. SP-D from both peaks bound to mannan-coated ELISA plates. SP-D from the high m.w. peak bound preferentially to intact influenza A virus and Gram-positive and Gram-negative bacteria, whereas the monomeric species preferentially bound to isolated LPS. Our data strongly suggest that polymorphic variation in the N-terminal domain of the SP-D molecule influences oligomerization, function, and the concentration of the molecule in serum. 相似文献
17.
辛纳毒蛋白是从香樟种子中分离的一种Ⅱ核糖体失活蛋白.最近,从香樟种子中还分离到另一种微型双链核糖体失活蛋白,命名为新丰毒蛋白.还原的新丰毒蛋白表现出与还原的辛纳毒蛋白同样的RNA N-糖苷酶和体外对抑制蛋白质翻译的活力.新丰毒蛋白的B链与辛纳毒蛋白的B链具有同样的分子质量和相同的N端10个氨基酸序列.它的A链N端10个氨基酸序列也与辛纳毒蛋白的A链完全一致,并且C端与辛纳毒蛋白的A链一样具有半胱氨酸,但是它的分子质量却只有辛纳毒蛋白A链的一半.RT-PCR和RNA印迹结果表明体内不存在新丰毒蛋白的mRNA.推测新丰毒蛋白是从辛纳毒蛋白通过蛋白质剪接而产生的,是一种研究蛋白质剪接的好材料. 相似文献
18.
《Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology》1989,994(3):195-199
The positions of the inter- and intra-chain disulfide bonds of human plasma α2HS-glycoprotein were determined. α2HS-glycoprotein was digested with acid proteinase and then with thermolysin. The disulfide bonds containing peptides were separated by reversed-phase HPLC and detected by SBD-F (7-fluorobenzo-2-oxa-1,3-diasole-4-sulfonic acid ammonium salt) method. One inter-disulfide bond containing peptide and five intra-disulfide bond containing peptides (A-chain) were purified and identified as Cys-18 (B-chain)-Cys-14 (A-chain), Cys-71-Cys-82, Cys-96-Cys-114, Cys-128-Cys-131, Cys-190-Cys-201 and Cys-212-Cys-229, respectively. The location of the intra-disulfide bonds revealed that the A-chain of α2HS-glycoprotein is composed of three domains. Two domains were shown to possess intramolecular homology judging from the total chain length of the domains, size of the loops formed by the SS bonds, the location of two disulfide loops near the C-terminal end of domains A and B, the distance between two SS bonds of each domain, the amino acid sequence homology between these two domains (22.6%), number of amino acid residues between the second SS loops and the end of domains A and B, and the positions of the ordered structures. 相似文献
19.
Articulatin-D, a 66 kDa ribosome inactivating protein (RIP) comprised of 29 kDa A-chain linked to 35 kDa B-chain, is purified from leafless mistletoe (Viscum articulatum) parasitic on Dalbergia sp. from Western Ghats (India). N-terminal sequence and LC-MS/MS analyses of A- and B-chain confirmed that articulatin-D is a type-2 RIP having high homology with other mistletoe lectins. Translation inhibition and diagnostic N-glycosidase activity of articulatin-D illustrate the presence of catalytically active A-chain. Its inability to: (i) bind to acid treated Sepharose CL-6B column, (ii) agglutinate trypsin-treated and untreated RBCs of human (A, B, O, AB), mice, rat, rabbit, buffalo, porcine, pigeon, cock, fish, sheep and goat even with 10 mg/ml of purified articulatin-D, (iii) show change in circular dichroism spectra after addition of sugar to the native protein, (iv) bind to different sugars (galactose, lactose, gal-NAc, rhamnose, arabinose, fucose and mannose) immobilized on Sepharose 4B matrix, and (v) show change in enthalpy during titration with galactose confirm that the B-chain of articulatin-D lacks sugar binding activity. Despite this, articulatin-D is highly toxic as characterized with low IC50 against different cancer cell lines (Jurkat: 0.31 ± 0.02 nM, MOLT-4: 0.51 ± 0.03 nM, U-937: 0.64 ± 0.07 nM, HL-60: 0.79 ± 0.11 nM, Raji: 1.45 ± 0.09 nM). Toxicity of RIPs has been ascribed to the absence/presence of B-chain with sugar binding activity. Identification of articulatin-D, the first cytotoxic RIP with B-chain lacking sugar binding activity opens new vistas in understanding cytotoxic action of RIPs. 相似文献
20.
C Panneerselvam M Clinton D Wellner B L Horecker 《Biochemical and biophysical research communications》1988,155(2):539-545
Parathymosin has been purified from calf liver and its primary sequence established, except for a segment containing approximately 11 amino acid residues in the central part of the polypeptide chain. Bovine parathymosin contains approximately 101 amino acid residues and shows 90% identity with rat parathymosin, with substitution of Glu for Asp at positions 21, 57, and 58, Asp for Glu at positions 60 and 63, and Ala for Val at position 77. Three non-conservative substitutions were Ala for Thr at position 81, Leu for Arg at position 78, and Val for Lys at position 79. The replacement at the last two positions of a pair of basic by hydrophobic amino acid residues may account for differences in chromatographic behavior observed for the bovine and rat polypeptides. Analysis of the NH2-terminus employed a new deblocking procedure which was also employed to analyze rat parathymosin, requiring correction of the previously published NH2-terminal sequence for that polypeptide. 相似文献