共查询到20条相似文献,搜索用时 31 毫秒
1.
N Smith A T Matheson M Yaguchi G E Willick R N Nazar 《European journal of biochemistry》1978,89(2):501-509
A 5-S RNA . protein complex has been isolated from the 50-S ribosomal subunit of an extreme halophile, Halobacterium cutirubrum. The 50-S ribosomal subunit from the extreme halophile requires 3.4 M K+ and 100 mM Mg2+ for stability. However, if the high K+ concentration is maintained but the Mg2+ concentration lowered to 0.3 mM, the 5-S RNA . protein complex is selectively extracted from the subunit. After being purified on an Agarose 0.5-m column the complex had a molecular weight of about 80000 and contained 5-S RNA and two proteins, HL13 and HL19, with molecular weights (by sedimentation equilibrium) of 18700 and 18000, respectively. No ATPase or GTPase activity could be detected in the 5-S RNA . protein complex. The amino acid composition and electrophoretic mobility on polyacrylamide gels indicated both proteins were much more acidic than the equivalent from Escherichia coli or Bacillus stearothermophilus. Partial amino acid sequence data suggest HL13 is homologous to EL18 and HL19 to EL5. 相似文献
2.
When Escherichia coli 30-S ribosomal subunits are hydrolysed under mild conditions, two ribonucleoprotein fragments of unequal size are produced. Knowledge of the RNA sequences contained in these hydrolysis products was required for the experiments described in the preceding paper, and the RNA sub-fragments have therefore been examined by oligonucleotide analysis. Two well-defined small fragments of free RNA, produced concomitantly with the ribonucleoprotein fragments, were also analysed. The larger ribonucleoprotein fragment, containing predominantly proteins S4, S5, S8, S15, S16 (17) and S20, contains a complex mixture of RNA sub-fragments varying from about 100 to 800 nucleotides in length. All these fragments arose from the 5'-terminal 900 nucleotides of 16-S RNA, corresponding to the well-known 12-S fragment. No long-range interactions could be detected within this RNA region in these experiments. The RNA from the smaller ribonucleoprotein fragment (containing proteins S7, S9 S10, S14 and S19) has been described in detail previously, and consists of about 450 nucleotides near the 3' end of the 16-S RNA, but lacking the 3'-terminal 150 nucleotides. The two small free RNA fragments (above) partly account for these missing 150 nucleotides; both fragments arose from section A of the 16-S RNA, but section J (the 3'-terminal 50 nucleotides) was not found. This result suggests that the 3' region of 16-S RNA is not involved in stable interactions with protein. 相似文献
3.
4.
Using immunological and chemical cleavage techniques, we have previously identified a domain contained within residues Phe56-Ser86 in the first tandem repeat (A1) of the heavy chain of factor XI which binds high Mr kininogen (Baglia, F. A., Jameson, B. A., and Walsh, P. N. (1990) J. Biol. Chem. 265, 4149-4154). We have now chemically synthesized peptides from corresponding homologous regions in the second (A2), third (A3), and fourth (A4) tandem repeats of the heavy chain (A2: Asn145-Ala176; A3: Asn235-Arg266; and A4: Gly326-Lys357). These peptides had no effect on the binding of factor XI to high Mr kininogen. Because of a lack of detailed structural information for the A1 domain, a molecular model of this region was constructed. This hypothetical model made distinct and testable predictions regarding potential surfaces and concomitant secondary structure. Specifically, the resulting structure depicted two juxtaposed beta-stranded stem-loops that, in conjunction with biological information, constitute a candidate surface for contact with high Mr kininogen. The hypothetical A1 model was, consequently, used as a predictive template in the rational design of two synthetic peptides (Val59-Arg70 and Asn72-Lys83). When both these peptides were added together and the binding of factor XI to high Mr kininogen was examined, a synergistic inhibitory effect was observed compared with each peptide added individually. Our data are consistent with the notion that the sequence of amino acids from Val59-Lys83 of the heavy chain of factor XI contains two antiparallel beta-strands connected by beta-turns that together comprise a continuous surface utilized for the binding of high Mr kininogen. 相似文献
5.
Tryptase from rat skin: purification and properties 总被引:2,自引:0,他引:2
Tryptase was purified 13,000-fold to apparent homogeneity from rat skin. The two-step procedure involved ammonium sulfate fractionation of the initial extract followed by combined sequential affinity chromatography on agarose-glycyl-glycyl-p-aminobenzamidine and concanavalin A-agarose. The purified enzyme had a specific activity toward N-benzoylarginine ethyl ester (BzArgOEt) of 170 mumol/min mg-1 and was obtained in a yield of 28% as determined by the specific substrate, H-D-Ile-Pro-Arg-p-nitroanilide. Rat skin tryptase was thermal labile, losing 50% of its activity when preincubated for 30 min at 30 degrees C. The presence of NaCl (1 M) improved thermal stability and was necessary for long-term storage. Heparin did not stabilize the enzyme against thermal denaturation, and heparin-agarose failed to bind the enzyme. Rat skin tryptase was inhibited by diisopropylphosphofluoridate, antipain, leupeptin, and aprotinin but not by alpha 1-antitrypsin, ovomucoid, or soybean or lima bean trypsin inhibitors. Substrate specificity studies using a series of tri- and tetrapeptidyl-p-nitroanilide and peptidyl-7-amino-4-methylcoumarin substrates demonstrated the existence of an extended substrate binding site. Rat skin tryptase hydrolyzed [Arg8]vasopressin, neurotensin, and the oxidized B-chain of insulin at the -Arg8-Gly9-NH2, -Arg8-Arg9-, and -Arg22-Gly23-bonds, respectively. No general proteinase activity was observed toward casein, hemoglobin, or azocoll. Rat skin tryptase had a Mr of 145,000 by gel filtration. The subunit Mr was either 34,000 or 30,000 depending on the electrophoretic technique used. Treatment of the enzyme with peptide N-glycosidase F (N-glycanase) decreased the subunit Mr by 4000. The enzyme exhibited multiple isoelectric forms (pI's of 4.5-4.9). Rat skin tryptase was found to be related statistically to other tryptases on the basis of amino acid composition. The N-terminal amino acid sequence was Ile1-Val2-Gly3-Gly4-Gln5-Glu6-Ala7-+ ++Ser8-Gly9-Asn10-Lys11-Trp12-Pro13- Trp14- Gln15-Val16-Ser17-Leu18-Arg19-Val20- --21-Asp-22Thr23-Tyr24-Typ25-, with a putative glycosylation site at residue 21. This sequence was 72-80% homologous with the N-terminus of other tryptases but only 40% homologous with that of bovine trypsin. 相似文献
6.
Bednarek MA Macneil T Tang R Fong TM Angeles Cabello M Maroto M Teran A 《Biopolymers》2008,89(5):401-408
alpha-Melanocyte stimulating hormone (alphaMSH), Ac-Ser(1)-Tyr(2)-Ser(3)-Met(4)-Glu(5)-His(6)-Phe(7)-Arg(8)-Trp(9)-Gly(10)-Lys(11)-Pro(12)-Val(13)-NH(2), is an endogenous agonist for the melanocortin receptor 1 (MC1R), the receptor found in the skin, several types of immune cells, and other peripheral sites. Three-dimensional models of complexes of this receptor with alphaMSH and its synthetic analog NDP-alphaMSH, Ac-Ser(1)-Tyr(2)-Ser(3)-Nle(4)-Glu(5)-His(6)-D-Phe(7)-Arg(8)-Trp(9)-Gly(10)-Lys(11)-Pro(12)-Val(13)-NH(2), have been previously proposed. In those models, the 6-9 segment of the ligand was considered essential for the ligand-receptor interactions. In this study, we probed the role of Trp(9) of NDP-alphaMSH in interactions with hMC1bR. Analogs of NDP-alphaMSH with various amino acids in place of Trp(9) were synthesized and tested in vitro in receptor affinity binding and cAMP functional assays at human melanocortin receptors 1b, 3, 4, and 5 (hMC1b,3-5R). Several new compounds displayed high agonist potency at hMC1bR (EC(50) = 0.5-5 nM) and receptor subtype selectivity greater than 2000-fold versus hMC3-5R. The Trp(9) residue of NDP-alphaMSH was determined to be not essential for molecular recognition at hMC1bR. 相似文献
7.
Sam68 RNA binding protein is an in vivo substrate for protein arginine N-methyltransferase 1 总被引:7,自引:0,他引:7
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)
Côté J Boisvert FM Boulanger MC Bedford MT Richard S 《Molecular biology of the cell》2003,14(1):274-287
RNA binding proteins often contain multiple arginine glycine repeats, a sequence that is frequently methylated by protein arginine methyltransferases. The role of this posttranslational modification in the life cycle of RNA binding proteins is not well understood. Herein, we report that Sam68, a heteronuclear ribonucleoprotein K homology domain containing RNA binding protein, associates with and is methylated in vivo by the protein arginine N-methyltransferase 1 (PRMT1). Sam68 contains asymmetrical dimethylarginines near its proline motif P3 as assessed by using a novel asymmetrical dimethylarginine-specific antibody and mass spectrometry. Deletion of the methylation sites and the use of methylase inhibitors resulted in Sam68 accumulation in the cytoplasm. Sam68 was also detected in the cytoplasm of PRMT1-deficient embryonic stem cells. Although the cellular function of Sam68 is unknown, it has been shown to export unspliced human immunodeficiency virus RNAs. Cells treated with methylase inhibitors prevented the ability of Sam68 to export unspliced human immunodeficiency virus RNAs. Other K homology domain RNA binding proteins, including SLM-1, SLM-2, QKI-5, GRP33, and heteronuclear ribonucleoprotein K were also methylated in vivo. These findings demonstrate that RNA binding proteins are in vivo substrates for PRMT1, and their methylation is essential for their proper localization and function. 相似文献
8.
Agustín Vioque Francisco Hernández Enrique Palacián 《Molecular and cellular biochemistry》1987,76(2):141-146
The contribution of lysine and arginine residues to the formation of yeast ribonucleoprotein complex 5S RNA. protein YL3 has been investigated by determining the effects on complex formation of modification with chemical reagents specific for either lysine or arginine. Treatment of protein YL3 with acetic anhydride, malefic anhydride or phenylglyoxal is accompanied by loss of its capacity to bind to 5S RNA. This effect is accomplished by modification with phenylglyoxal of only 3 arginine residues per YL3 molecule. In contrast, a large number of protein YL3 amino groups [16] must be modified by acetic anhydride to prevent complex formation. 相似文献
9.
Cell-adhesion molecules are critical for immune response. It is well known that the inhibition of adhesion is very effective in immunotherapy and that the peptides derived from leukocyte function associated antigen (LFA-1) and intercellular adhesion molecule (ICAM-1) modulate cell-adhesion interaction. The three-dimensional structure of a cyclic peptide, Cyclo(1,12)Pen(1)-Asp(2)-Leu(3)-Ser(4)-Tyr(5)-Ser(6)-Leu(7)-Asp(8)-Asp(9)-Leu(10)-Arg(11)-Cys(12) (cLBEL) derived from the beta subunit of LFA-1 which is known to modulate homotypic T-cell-adhesion process has been studied using NMR, CD and molecular dynamics (MD) simulation. The peptide exhibits two possible conformations in solution. Structure I has a conformation with two consecutive beta-turns involving residues Tyr(5)-Ser(6)-Leu(7)-Asp(8) and Asp(9)-Leu(10)-Arg(11)-Cys(12). Structure II has a beta-turn at Tyr(5)-Ser(6)-Leu(7)-Asp(8) and forms a beta-hairpin type of conformation. 相似文献
10.
Two peptic fragments (residues 37-88 and 43-88) of guinea pig myelin basic protein which are capable of inducing experimental allergic encephalomyelitis in Lewis rats were cleaved to shorter fragments with alpha-protease (Crotalus atrox proteinase, EC 3.4.24.1) and thermolysin (EC 3.4.24.4). The fragments were isolated, purified, and identified by amino acid composition and NH2- and COOH-terminal residues. The time courses of the reactions, monitored by thin layer electrophoresis of the digests, showed that alpha-protease cleaves peptide (43-88) initially at the Pro(71)-Gln(72) bond, and that the product peptides are subsequently attacked at the Arg(63) -Thr(64), Ser(74)-Gln(75), Arg(78)-Ser(79), and Ser(76)-Gln(80) bonds. No significant cleavages occurred at the -Leu, -Val, and -Ala bonds. These results are in striking contrast to those obtained previously by others workers with other peptide substrates, where selective cleavage at hydrophobic residues occurred. Thermolysin was found to attack peptide (37-88) at the Phe(42)-Phe(43) bond very rapidly; the product peptides were subsequently attacked at the His(60)-Ala(61), Ser(38)-Ile(39)-Tyr(67)-Gly(68), and Pro(84)-Val(85) bonds. These cleavages are compatible with the known specificity of this enzyme. Several of the fragments prepared with these two enzymes, peptides (43-71), (61-88), (75-88), and (72-84) have been used in other studies to locate the encephalitogenic site in the parent peptic peptide. 相似文献
11.
I Winicov 《Biochimica et biophysica acta》1975,402(1):62-68
The synthesis of 5-S RNA was found to be unchanged at both the permissive (33.5 degrees C) and non-permissive (38.5 degrees C) temperatures in a temperature-sensitive Baby Hamster Kidney cell line (BHK 21 ts 422 E) as measured relative to synthesis of 18-S rRNA. The 5-S RNA is shown to be associated with nucleolar ribonucleoprotein particles even though rRNA processing does not yield a functional 28-S rRNA at the non-permissive temperature. The amount of 5-S RNA found associated with the 80-S ribonucleoprotein particles was the same at the permissive and non-permissive temperatures, indicating that an aberrant 5-S RNA contribution to rRNA processing is not a primary cause for the temperature-sensitive lesion of rRNA maturation in this mutant cell line. The amount of 5-S RNA in nucleolar 80-S RNA particles indicated that the association of 5-S RNA with the rRNA precursor particle occurs before the cleavage step at which 32-S precursor RNA is produced. 相似文献
12.
Aspartate transcarbamylase (EC 2.1.3.2) from E. coli is a multimeric enzyme consisting of two catalytic subunits and three regulatory subunits whose activity is regulated by subunit interactions. Differential scanning calorimetric (DSC) scans of the wild-type enzyme consist of two peaks, each comprised of at least two components, corresponding to denaturation of the catalytic and regulatory subunits within the intact holoenzyme (Vickers et al., J. Biol. Chem. 253 (1978) 8493; Edge et al., Biochemistry 27 (1988) 8081). We have examined the effects of nine single-site mutations in the catalytic chains. Three of the mutations (Asp-100-Gly, Glu-86-Gln, and Arg-269-Gly) are at sites at the C1: C2 interface between c chains within the catalytic subunit. These mutations disrupt salt linkages present in both the T and R states of the molecule (Honzatko et al., J. Mol. Biol. 160 (1982) 219; Krause et al., J. Mol. Biol. 193 (1987) 527). The remainder (Lys-164-Ile, Tyr-165-Phe, Glu-239-Gln, Glu-239-Ala, Tyr-240-Phe and Asp-271-Ser) are at the C1: C4 interface between catalytic subunits and are involved in interactions which stabilize either the T or R state. DSC scans of all of the mutants except Asp-100-Gly and Arg-269-Gly consisted of two peaks. At intermediate concentrations, Asp-100-Gly and Arg-269-Gly had only a single peak near the Tm of the regulatory subunit transition in the holoenzyme, although their denaturational profiles were more complex at high and low protein concentrations. The catalytic subunits of Glu-86-Gln, Lys-164-Ile and Asp-271-Ser appear to be significantly destabilized relative to wild-type protein while Tyr-165-Phe and Tyr-240-Phe appear to be stabilized. Values of delta delta G degree cr, the difference between the subunit interaction energy of wild-type and mutant proteins, evaluated as suggested by Brandts et al. (Biochemistry 28 (1989) 8588) range from -3.7 kcal mol-1 for Glu-86-Gln to 2.4 kcal mol-1 for Tyr-165-Phe. 相似文献
13.
T Ekida C Nishimura S Masuda S Itoh I Shimada Y Arata 《Biochemical and biophysical research communications》1992,189(1):211-220
In order to locate the receptor-binding region of human interleukin-6 (IL-6), twelve peptide fragments were prepared by digestion of IL-6 with lysylendopeptidase. A significant activity of the receptor-binding was observed only for a peptide Ile88-Lys121, although the activity was estimated at 10(4)-fold less than that of intact IL-6. Solution structure of the peptide Ile88-Lys121 was analyzed by using two-dimensional nuclear magnetic resonance (NMR) spectroscopy. The results indicate the presence of alpha-helices in the regions Leu93-Phe106 and Glu110-Ser119. On the basis of the NMR data, we also prepared two peptides. Four-fold less binding activity than that of the peptide Ile88-Lys121 was observed for the peptide Ile88-Arg105, but no activity for the peptide Glu110-Lys121. These results suggest that the helical peptide Ile88-Arg105 composes a part of the receptor-binding region. 相似文献
14.
Andreas G Tzakos Patrick Fuchs Nico A J van Nuland Anastasios Troganis Theodore Tselios Spyros Deraos John Matsoukas Ioannis P Gerothanassis Alexandre M J J Bonvin 《European journal of biochemistry》2004,271(16):3399-3413
Experimental autoimmune encephalomyelitis can be induced in susceptible animals by immunodominant determinants of myelin basic protein (MBP). To characterize the molecular features of antigenic sites important for designing experimental autoimmune encephalomyelitis suppressing molecules, we report structural studies, based on NMR experimental data in conjunction with molecular dynamic simulations, of the potent linear dodecapeptide epitope of guinea pig MBP, Gln74-Lys75-Ser76-Gln77-Arg78-Ser79-Gln80-Asp81-Glu82-Asn83-Pro84-Val85 [MBP(74-85)], and its antagonist analogue Ala81MBP(74-85). The two peptides were studied in both water and Me(2)SO in order to mimic solvent-dependent structural changes in MBP. The agonist MBP(74-85) adopts a compact conformation because of electrostatic interactions of Arg78 with the side chains of Asp81 and Glu82. Arg78 is 'locked' in a well-defined conformation, perpendicular to the peptide backbone which is practically solvent independent. These electrostatic interactions are, however, absent from the antagonist Ala81MBP(74-85), resulting in great flexibility of the side chain of Arg78. Sequence alignment of the two analogues with several species of MBP suggests a critical role for the positively charged residue Arg78, firstly, in the stabilization of the local microdomains (epitopes) of the integral protein, and secondly, in a number of post-translational modifications relevant to multiple sclerosis, such as the conversion of charged arginine residues to uncharged citrullines. Flexible docking calculations on the binding of the MBP(74-85) antigen to the MHC class II receptor site I-A(u) using haddock indicate that Gln74, Ser76 and Ser79 are MHC II anchor residues. Lys75, Arg78 and Asp81 are prominent, solvent-exposed residues and, thus, may be of importance in the formation of the trimolecular T-cell receptor-MBP(74-85)-MHC II complex. 相似文献
15.
Alpha-melanotropin (alphaMSH), Ac-Ser1-Tyr2-Ser3-Met4-Glu5-His6-Phe7-Arg8-Trp9-Gly10-Lys11-Pro12-Val13-NH2,(1) has been long recognized as an important physiological regulator of skin and hair pigmentation in mammals. Binding of this peptide to the melanocortin receptor 1 (MC1R) leads to activation of tyrosinase, the key enzyme of the melanin biosynthesis pathway. In this study, interactions of the human MC1bR (an isoform of the receptor 1a) with the synthetic cyclic analogs of alphaMSH were studied. These ligands were analogs of MTII, Ac-Nle4-cyclo-(Asp5-His6-D-Phe7-Arg8-Trp9-Lys10)-NH2, a potent pan-agonist at the human melanocortin receptors (hMC1,3-5R). In the structure of MTII, the His6-D-Phe7-Arg8-Trp9 segment has been recognized as "essential" for molecular recognition at the human melanocortin receptors (hMC1,3-5R). Herein, the role of the Trp9 in the ligand interactions with the hMC1b,3-5R has been reevaluated. Analogs with various amino acids in place of Trp9 were synthesized and tested in vitro in receptor affinity binding and cAMP functional assays at human melanocortin receptors 1b, 3, 4 and 5 (hMC1b,3-5R). Several of the new peptides were high potency agonists (partial) at hMC1bR (EC50 from 0.5 to 20 nM) and largely inactive at hMC3-5R. The bulky aromatic side chain in position 9, such as that in Trp, was found not to be essential to agonism (partial) of the studied peptides at hMC1bR. 相似文献
16.
17.
The spatial structure of the cardioactive octapeptide Pro1-Gln2-Asp3-Pro4-Phe5-Leu6-Arg7-Ile8-NH2 was investigated using the theoretical conformational analysis. The low-energy conformations of the octapeptide molecule were found, the values of dihedral angles of the backbone and side chains of the amino acid residues constituting the peptide were determined, and the energies of intra-and interresidual interactions were estimated. It was shown that the spatial structure of this molecule represent six stable low-energy forms of the main chain. 相似文献
18.
J Garin L Michel A Dupuis J P Issartel J Lunardi J Hoppe P Vignais 《Biochemistry》1989,28(4):1442-1448
[32P]Azidonitrophenyl phosphate [( 32P]ANPP) is a photoactivatable analogue of Pi. It competes efficiently with Pi for binding to the F1 sector of beef heart mitochondrial ATPase and photolabels the Pi binding site located in the beta subunit of F1 [Lauquin, G. J. M., Pougeois, R., & Vignais, P. V. (1980) Biochemistry 19, 4620-4626]. By cleavage of the photolabeled beta subunit of F1 with cyanogen bromide, trypsin, and chymotrypsin, bound [32P]ANPP was localized in a fragment spanning Thr 299-Phe 326. By Edman degradation of the radiolabeled tryptic peptide spanning Ile 296-Arg 337, [32P]ANPP was found to be attached covalently by its photoreactive group to Ile 304, Gln 308, and Tyr 311. These results are discussed in terms of a model in which the phosphate group of [32P]ANPP interacts with a glycine-rich sequence of the beta subunit, spanning Gly 156-Lys 162, which is spatially close to the photolabeled Ile 304-Tyr 311 segment of the same subunit. 相似文献
19.
The complete nucleotide sequence of 5-S RNA from Bacillus licheniformis was determined by analysis of complete and partial digests obtained with either T1 or pancreatic ribonuclease. The molecule was found to have a length of 116 nucleotides and may possess a minor sequence heterogeneity. There is a large degree of homology between the sequence of B. licheniformis 5-S RNA and those published for 5-S RNA from B. megatherium and B. stearothermophilus. The difference between the three 5-S RNA species are limited mainly to the two terminal and one internal sequence. B. licheniformis 5-S RNA contains the sequence U95-G-A-G-A-G100, which in B. subtilis has been implicated in the processing of precursor 5-S RNA. Possible models for the secondary structure of prokaryotic 5-S RNA are discussed on the basis of the results of limited digestion of B. licheniformis 5-S RNA by ribonuclease T1. 相似文献
20.
Mzareulov KD 《Biofizika》2005,50(3):413-417
The method of theoretical conformational analysis was used to study the inverse structural problem to determine the amino acid sequence of the peptide molecule capable of inhibiting the site of binding of p21 to cell receptors. At the first stage of the computational experiment, the spatial structure and the conformational possibilities of the binding sites of protein p21 and its cellular receptors were determined. Then the three-dimensional structures of several peptides containing the Arg-Ala-Ala-Glu-Asp site were studied. By varying the number of alanine residues in the adjacent regions of the molecule, the sequence H-Asp1-Ala2-Ala3-Ala4-Arg5-Ala6-Ala7-Glu8-Asp9-Ala10-Ala11--Lys12-QH was chosen, which most adequately simulates the conformational properties of the address fragments of oncoprotein receptors. The peptide-molecule having this primary structure is capable of forming a complex with p21, i.e., blocking the binding site of the oncoprotein by preventing the signal transduction from the oncoprotein to the cell, thereby breaking the cycle of the carcinogenic process. 相似文献