共查询到20条相似文献,搜索用时 15 毫秒
1.
K J Nielsen P R Barron D J Craik P F Lambert J D Wade 《Biochemistry international》1991,25(2):199-210
Two dimensional NMR spectra of porcine brain natriuretic peptide have been recorded at 400 MHz. Peak assignments have been made and the combined information from chemical shifts, coupling constants, temperature coefficients, exchange studies and nuclear Overhauser effects has been used to determine the conformation of pBNP in aqueous media. Overall the peptide appears to be conformationally averaged with the possibility of some restricted flexibility in localized regions. The conformation of porcine brain natriuretic peptide in water is compared to previous studies in d6-DMSO and to studies of atrial natriuretic peptide and some closely related analogues in H2O and d6-DMSO. 相似文献
2.
Two-dimensional 1H NMR studies of histidine-containing protein from Escherichia coli. 1. Sequential resonance assignments 总被引:4,自引:0,他引:4
Two-dimensional NMR studies at 500 MHz have been performed on the histidine-containing protein (HPr) from Escherichia coli. HPr is one of the phosphocarrier proteins involved in the bacterial phosphoenolpyruvate:sugar phosphotransferase system that is responsible for the concomitant phosphorylation and translocation of a number of sugars. Sequential resonance assignments of HPr are complete. The conventional method of sequential assignments involving J-correlated spectroscopy (COSY) and nuclear Overhauser spectroscopy (NOESY) has been supplemented by optimized relayed coherence transfer spectroscopy (RELAY) to help overcome the spectral overlap that is inevitable in the spectra of proteins the size of HPr. RELAY experiments were performed in H2O to obtain NH-C beta H connectivities and in D2O to obtain C alpha H-C gamma H connectivities. The abundance of relayed coherence transfer peaks in the two experiments greatly aided in the assignment process of the complicated protein spectrum. The assignments lay the groundwork for the determination of the solution structure of HPr, as described in the accompanying paper [Klevit, R. E., & Waygood, E. B. (1986) Biochemistry (third paper of three in this issue)]. 相似文献
3.
Two enzymatically modified derivatives of tobramycin have been prepared by gentamicin nucleotidyl transferase-catalyzed adenylylation of tobramycin, using ATP and (Sp)-ATP alpha S as adenylylation substrates. (EC 2.7.7.46). The 1H, 13C, and 31P NMR spectra have been assigned for tobramycin, 2'-(adenosine-5'-phosphoryl)-tobramycin (TbAMP) and 2'-(adenosine-5'-thiophosphoryl)-tobramycin (TbAMPS). Several one- and two-dimensional NMR techniques have been utilized, notably, 1H-1H homonuclear correlation spectroscopy at 470 or 500 MHz and 13C-1H heteronuclear correlation spectroscopy at 50.3 MHz. The 1H assignments for tobramycin are similar to those previously reported for rings I and III of kanamycin A. The 13C assignments for tobramycin were similar to those previously reported, except for reversal of the assignments for anomeric carbons in the glycosyl rings. The 1H and 13C assignments for tobramycin were used to guide the assignments of the spectra for TbAMP and TbAMPS. Nearly complete assignments were obtained for these two derivatives of tobramycin. From the measured proton coupling constants, only small conformational changes were observed upon modification of tobramycin by adenylylation. From the proton and carbon spectra of the adenylylated derivatives the 2' position is shown to be the site of adenylation. Large downfield shifts of the 2'proton and carbon resonances are easily observed and are more pronounced for TbAMPS than for TbAMP. 相似文献
4.
Coupal M De Léan A McNicoll N Fournier A 《Biochemical and biophysical research communications》1999,258(1):81-86
Two Nalpha-benzophenone-substituted photoprobes, derived from the high affinity NPR-A chimeric agonist [N, C, rANP(1-28)]pBNP32 (pBNP1) were assembled by solid-phase peptide synthesis. [Nalpha-p-benzoylbenzoyl, Tyr2]pBNP1 (probe A), and [Nalpha-p-benzoylbenzoyl, Tyr18]pBNP1 (probe B) were synthesized and their affinity was tested on bovine zona glomerulosa membrane preparations. Both were found to exert ANP-type high affinities (Kd = 20 pM) with Kd of 10 pM and 30 pM for probe A, and probe B, respectively. Photolabeling of NPR-A with both analogs cross-linked specifically the 130 kDa monomeric NPR-A. The maximal irreversible ligand incorporations were estimated at 18% and 41% for probe A, and probe B, respectively. These results show that the N-terminus of the chimeric compound can be acylated with a large chemical function, such as the benzophenone moiety, without loosing its affinity for the NPR-A receptor. Furthermore, Leu2 or Leu18 can be substituted with tyrosine without disturbing the binding capacity of the ligand. Finally, it appears that the pBNP1 N-terminus is close to the receptor structure as irreversible incorporation is observed after photolabeling. 相似文献
5.
M Petersheim R L Moldow H N Halladay A J Kastin A J Fischman 《International journal of peptide and protein research》1992,40(1):41-48
N-Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2) is an endogenous brain peptide with multiple effects on animal behavior. However, there have been no studies on the conformation of this tetrapeptide. In this report, we studied the conformation of N-Tyr-MIF-1 in aqueous solution by conventional one-dimensional and two-dimensional (COSY and NOESY) 1H nuclear magnetic resonance spectroscopy at 300 MHz. A complete set of assignments for the resolved resonances and approximate assignments for the overlapping resonances were made. The results demonstrate that N-Tyr-MIF-1 is in slow exchange between two conformers, most likely determined by the cis and trans states of the proline residue. The minor conformation represents 30 +/- 3% of the population over the temperature range from 3 degrees to 73 degrees. In the major conformation, the tyrosine aromatic ring appears to be close enough to interact directly with the proline pyrrolidine ring, as indicated by a strong temperature dependence of the proline C beta H, C delta H and C delta H' chemical shifts. In contrast, this interaction of the tyrosine and proline rings is not present in the minor conformation. 相似文献
6.
Two synthetic peptides from the beta 2 subunit of tryptophan synthase have been studied by 1H-NMR spectroscopy at 300 MHz. One peptide, His-Gly-Arg-Val-Gly-Ile-Tyr-Phe-Gly-Met-Lys (peptide 11; Ile, isoleucine) is antigenic and binds with a high affinity to a monoclonal antibody that recognizes the native beta 2 subunit. The second peptide, His-Gly-Arg-Val-Gly-Ile-Tyr-Phe (peptide 8) reacts very weakly with the antibody. The 1H-NMR spectra of the two peptides have been assigned from two-dimensional techniques in H2O, 2H2O and (2H6) dimethyl sulfoxide [(2H6)Me2SO]. The structure has been evaluated through analysis of nuclear Overhauser effects, coupling constants, amide-proton exchange rates and their temperature coefficients, and chemical shifts. In aqueous solvent, the C-terminal part of peptide 11 presents some structure centered around residues Phe-Gly-Met. The relationship between the structure found in peptide 11 and its antigenic nature is discussed. 相似文献
7.
G Otting W E Steinmetz P E Bougis H Rochat K Wüthrich 《European journal of biochemistry》1987,168(3):609-620
Sequence-specific assignments of the 1H-nuclear magnetic resonance (NMR) spectra of the cardiotoxins CTXIIa and CTXIIb from Naja mossambica mossambica were obtained using two-dimensional NMR experiments at 500 MHz and the independently determined amino acid sequences. Assignments were obtained from data at 25 degrees C and 45 degrees C for all but one backbone proton of the 60 residues in each protein. Complete or partial assignments are also reported for the side-chain protons. These assignments supercede those published previously for the toxin preparation VII2 [Hosur, R. V., Wider, G. & Wüthrich K. (1983) Eur. J. Biochem. 130, 497-508]. The 1H/2H-exchange kinetics were measured in 2H2O at 20 degrees C for the amide protons and the N-terminal amino group. These and additional NMR data enabled the determination of the secondary structure in aqueous solution, which is virtually identical in CTXIIa and CTXIIb. Both proteins contain a short double-stranded antiparallel beta-sheet comprising the residues 2-4 and 11-13, and a triple-stranded antiparallel beta-sheet consisting of the residues 20-26, 35-39, and 49-55. The two peripheral strands of the triple-stranded beta-structure were found to be connected by a right-handed cross-over, and the locations of several tight turns were also identified. 相似文献
8.
9.
1H, 13C, and 15N backbone assignment and secondary structure of the receptor-binding domain of vascular endothelial growth factor. 总被引:1,自引:0,他引:1
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W. J. Fairbrother M. A. Champe H. W. Christinger B. A. Keyt M. A. Starovasnik 《Protein science : a publication of the Protein Society》1997,6(10):2250-2260
Nearly complete sequence-specific 1H, 13C, and 15N resonance assignments are reported for the backbone atoms of the receptor-binding domain of vascular endothelial growth factor (VEGF), a 23-kDa homodimeric protein that is a major regulator of both normal and pathological angiogenesis. The assignment strategy relied on the use of seven 3D triple-resonance experiments [HN(CO)CA, HNCA, HNCO, (HCA)CONH, HN(COCA)HA, HN(CA)HA, and CBCA-(CO)NH] and a 3D 15N-TOCSY-HSQC experiment recorded on a 0.5 mM (12 mg/mL) sample at 500 MHz, pH 7.0, 45 degrees C. Under these conditions, 15N relaxation data show that the protein has a rotational correlation time of 15.0 ns. Despite this unusually long correlation time, assignments were obtained for 94 of the 99 residues; 8 residues lack amide 1H and 15N assignments, presumably due to rapid exchange of the amide 1H with solvent under the experimental conditions used. The secondary structure of the protein was deduced from the chemical shift indices of the 1H alpha, 13C alpha, 13C beta, and 13CO nuclei, and from analysis of backbone NOEs observed in a 3D 15N-NOESY-HSQC spectrum. Two helices and a significant amount of beta-sheet structure were identified, in general agreement with the secondary structure found in a recently determined crystal structure of a similar VEGF construct [Muller YA et al., 1997, Proc Natl Acad Sci USA 94:7192-7197]. 相似文献
10.
Sequential 1H NMR assignments and secondary structure of the kringle domain from urokinase. 总被引:1,自引:0,他引:1
The sequence-specific 1H NMR assignments of the 89-residue recombinant kringle domain from human urokinase are presented. These were achieved primarily by utilizing TOCSY and NOESY spectra in conjunction with COSY spectra recorded at 500 MHz and 600 MHz. Regular secondary structure elements have been derived from a qualitative interpretation of nuclear Overhauser enhancement, JNH alpha coupling constant, and amide proton exchange data. Two helices have been identified. One helix, involving Ser40-Gly46, corresponds to that reported for t-PA kringle 2 (Byeon et al., 1991), but does not exist in other kringles with known structures. The second helix, in the region Asn26-Gln33, is thus far unique to the urokinase kringle. Three antiparallel beta-sheets and three tight turns have also been identified, which correspond exactly to those identified in t-PA kringle 2 both in solution and in the crystalline state (de Vos et al., 1992). Despite the very different ligand binding properties of the urokinase kringle, NOE data indicate that the tertiary fold of the molecule conforms closely to that found for other kringles. 相似文献
11.
A recombinant 90-residue polypeptide fragment containing the three-loop kringle-2 domain of human tissue-type plasminogen activator (t-PA) has been studied by two-dimensional 1H-NMR spectroscopy at 500 MHz. Complete sequence-specific resonance assignments were derived. Overall, the kringle exhibits a compact, folded conformation with more than 50% of the residues in irregular structures. Elements of secondary structure were identified from sequential, medium- and long-range dipolar (Overhauser) interproton interactions. These identifications were corroborated by analysis of spin-spin scalar 3J alpha N splittings and identification of backbone amide NH protons exhibiting retarded 1H/2H exchange in 2H2O. Three antiparallel beta-sheets and six tight turns were located. In addition, one short alpha-helical region was found in the Ser43-Ala44-Gln44a-Ala44b-Leu44c-Gly45+ ++ segment; this region contains three-residue insertions unique to the t-PA and urokinase kringles. Although the secondary structure of the t-PA kringle 2 in solution is in overall agreement with that observed in the crystallographic structure of the prothrombin kringle 1 [Tulinsky, A., Park, C.H. & Skrzypczak-Jankun, E. (1988) J. Mol. Biol. 202, 885-901], the alpha-helical segment and other details of the secondary structure differ somewhat from the prothrombin homolog. 相似文献
12.
Summary The ultracytochemical localization of particulate guanylate cyclase has been studied in lamb olfactory mucosa after activation with rat atrial natriuretic factor (rANF), porcine brain natriuretic peptide (pBNP), porcine C-type natriuretic peptide (pCNP) or rat brain natriuretic peptide (rBNP). Particulate guanylate cyclase is the receptor for these peptides and recently two subtypes of the cyclase have been identified. These isoforms are stimulated differently by ANF, BNP and CNP. Under our experimental conditions, rANF, pCNP and pBNP were strong activators of particulate guanylate cyclase in lamb olfactory mucosa, as demonstrated by the presence of reaction product. Samples incubated in basal conditions without rANF, pCNP or pBNP, or samples incubated in presence of rBNP did not reveal any cyclase activity. The rANF-stimulated cyclase activity was localized in the apical portion of olfactory epithelium. pCNP-stimulated guanylate cyclase was detected to the lamina propria in association with secretory cells of Bowman's glands and with cells in close relation with Bowman's glands (elongated cells and myoepithelial cells). The cyclase activity stimulated by pBNP was limited to cells of Bowman's glands. The present data indicate that ANF and CNP are recognized by different receptors and that BNP and CNP bind to the same receptor. 相似文献
13.
IIIGlc is an 18.1-kDa signal-transducing phosphocarrier protein of the phosphoenolpyruvate:glycose phosphotransferase system (PTS) of Escherichia coli. Virtually complete (98%) backbone 1H, 15N, and 13C nuclear magnetic resonance (NMR) signal assignments were determined by using a battery of triple-resonance three-dimensional (3D) NMR pulse sequences. In addition, nearly complete (1H, 95%; 13C, 85%) side-chain 1H and 13C signal assignments were obtained from an analysis of 3D 13C HCCH-COSY and HCCH-TOCSY spectra. These experiments rely almost exclusively upon one- and two-bond J couplings to transfer magnetization and to correlate proton and heteronuclear NMR signals. Hence, essentially complete signal assignments of this 168-residue protein were made without any assumptions regarding secondary structure and without the aid of a crystal structure, which is not yet available. Moreover, only three samples, one uniformly 15N-enriched, one uniformly 15N/13C-enriched, and one containing a few types of amino acids labeled with 15N and/or 13C, were needed to make the assignments. The backbone assignments together with the 3D 15N NOESY-HMQC and 13C NOESY-HMQC data have provided extensive information about the secondary structure of this protein [Pelton, J.G., Torchia, D.A., Meadow, N.D., Wong, C.-Y., & Roseman, S (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 3479-3488]. The nearly complete set of backbone and side-chain atom assignments reported herein provide a basis for studies of the three-dimensional structure and dynamics of IIIGlc as well as its interactions with a variety of membrane and cytoplasmic proteins. 相似文献
14.
Sunilkumar PN Sadasivan C Devaky KS Haridas M 《Indian journal of biochemistry & biophysics》2007,44(1):44-49
Kaliocin-1, a 31-residue synthetic peptide (FFSASCVPGADKGQFPNLCRLCA GTGENKCA), which has shown the antimicrobial activity forms the 152-182 fragment of human lactoferrin (HLf). As the octapeptide FSASCVPG forms the 2-9 fragment of kaliocin-1, in the present study, its conformation in dimethyl sulfoxide-d6 (DMSO-d6) has been determined using two-dimensional (2D) nuclear magnetic resonance (NMR) spectroscopy as well as restrained molecular dynamics. Sequence specific assignments of all the 1H resonances have been carried out using 2D correlation experiments (2D DQF-COSY, TOCSY and ROESY). In dimethyl sulfoxide-d6 at 25 degrees C, the octapeptide adopts a predominantly extended backbone conformation. The calculated structure resembles closely with the reported structure of the corresponding fragment of HLf. The peptide also has sequence and structural similarity with the corresponding fragments of lactoferrins from other organisms. 相似文献
15.
Proton electron-nuclear double-resonance spectra of molybdenum(V) in different reduced forms of xanthine oxidase 总被引:1,自引:0,他引:1
B D Howes N M Pinhal N A Turner R C Bray G Anger A Ehrenberg J B Raynor D J Lowe 《Biochemistry》1990,29(26):6120-6127
Electron-nuclear double-resonance (ENDOR) spectra of protons coupled to molybdenum(V) in reduced xanthine oxidase samples have been recorded. Under appropriate conditions these protons may be studied without interference from protons coupled to reduced iron-sulfur centers. Spectra have been obtained for the molybdenum(V) species known as Rapid, Slow, Inhibited, and Desulfo Inhibited. Resonances corresponding to at least nine protons or sets of protons are observed for all four species, with coupling constants in the range 0.08-4 MHz. Most of these protons do not exchange when 2H2O is used as solvent. Additional protons giving couplings up to 40 MHz are also detected. These correspond to EPR-detectable protons studied in earlier work. The strongly coupled protons may be replaced by 2H, through appropriate use of 2H2O or of 2H-substituted substrates, with consequent disappearance of the 1H resonances. In most cases the corresponding 2H ENDOR features have also been observed. The nature of the various coupled protons is briefly discussed. Results permit specific conclusions to be drawn about the structures of the Inhibited and Desulfo Inhibited species. In particular, the data indicate that the aldehyde residue of the Inhibited species has been oxidized and that the four protons derived from the ethylene glycol molecule in the Desulfo Inhibited species are not all equivalent. Recent assignments [Edmondson, D.E., & D'Ardenne, S.C. (1989) Biochemistry 28, 5924-5930] of the weakly coupled protons in the latter species appear not to be soundly based. The possibility of obtaining more detailed structural information from the spectra is briefly considered.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
16.
Katharine A. Carpenter Brian C. Wilkes Andr De Lan Alain Fournier Peter W. Schiller 《Biopolymers》1997,42(1):37-48
A conformational study by nmr spectroscopy was performed with the highly active 28-residue hybrid natriuretic peptide analogue pBNP1 [M. Mimeault, A. De Léan, M. Lafleur, D. Bonenfant, and A. Fournier (1995) Biochemistry, Vol. 34, pp. 955–964], which consists of the cyclic peptide core of pBNP32 and the N- and C-terminal exocyclic segments of rANP(99–126). In purely aqueous solution pBNP1 exhibits random coil behavior as evidenced by the almost complete absence of structurally significant nmr observables. By contrast, elements of secondary structure emerged upon the addition of dodecylphosphocholine micelles to the aqueous sample. Nuclear Overhauser effect distance-restrained molecular dynamics simulations in conjunction with torsional angle determinations permitted the generation of a reasonable model of the lipid-bound conformation of pBNP1. According to this model, pBNP1 adopts turn-like features in the cyclic and C-terminal regions of the peptide, but remains quite flexible in the N-terminal segment. Two hydrophobic cores separated by a hydrophilic cleft were also evident in the generated structure. A mechanism is proposed whereby the hydrophobic interactions necessary to stabilize a folded structure of pBNP1 are facilitated by the presence of the membrane-like polar/apolar interface provided by the phospholipid micelles. © 1997 John Wiley & Sons, Biopoly 42: 37–48, 1997 相似文献
17.
Multinuclear two-dimensional NMR techniques were used to assign nearly all diamagnetic 13C and 15N resonances of the plant-type 2Fe.2S* ferredoxin from Anabaena sp. strain PCC 7120. Since a 13C spin system directed strategy had been used to identify the 1H spin systems [Oh, B.-H., Westler, W. M., & Markley, J. L. (1989) J. Am. Chem. Soc. 111, 3083-3085], the sequence-specific 1H assignments [Oh, B.-H., & Markley, J. L. (1990) Biochemistry (first paper of three in this issue)] also provided sequence-specific 13C assignments. Several resonances from 1H-13C groups were assigned independently of the 1H assignments by considering the distances between these nuclei and the paramagnetic 2Fe.2S* center. A 13C-15N correlation data set was used to assign additional carbonyl carbons and to analyze overlapping regions of the 13C-13C correlation spectrum. Sequence-specific assignments of backbone and side-chain nitrogens were based on 1H-15N and 13C-15N correlations obtained from various two-dimensional NMR experiments. 相似文献
18.
W J Fairbrother A G Palmer M Rance J Reizer M H Saier P E Wright 《Biochemistry》1992,31(18):4413-4425
Nearly complete assignment of the aliphatic 1H and 13C resonances of the IIAglc domain of Bacillus subtilis has been achieved using a combination of double- and triple-resonance three-dimensional (3D) NMR experiments. A constant-time 3D triple-resonance HCA(CO)N experiment, which correlates the 1H alpha and 13C alpha chemical shifts of one residue with the amide 15N chemical shift of the following residue, was used to obtain sequence-specific assignments of the 13C alpha resonances. The 1H alpha and amide 15N chemical shifts had been sequentially assigned previously using principally 3D 1H-15N NOESY-HMQC and TOCSY-HMQC experiments [Fairbrother, W. J., Cavanagh, J., Dyson, H. J., Palmer, A. G., III, Sutrina, S. L., Reizer, J., Saier, M. H., Jr., & Wright, P. E. (1991) Biochemistry 30, 6896-6907]. The side-chain spin systems were identified using 3D HCCH-COSY and HCCH-TOCSY spectra and were assigned sequentially on the basis of their 1H alpha and 13C alpha chemical shifts. The 3D HCCH and HCA(CO)N experiments rely on large heteronuclear one-bond J couplings for coherence transfers and therefore offer a considerable advantage over conventional 1H-1H correlation experiments that rely on 1H-1H 3J couplings, which, for proteins the size of IIAglc (17.4 kDa), may be significantly smaller than the 1H line widths. The assignments reported herein are essential for the determination of the high-resolution solution structure of the IIAglc domain of B. subtilis using 3D and 4D heteronuclear edited NOESY experiments; these assignments have been used to analyze 3D 1H-15N NOESY-HMQC and 1H-13C NOESY-HSQC spectra and calculate a low-resolution structure [Fairbrother, W. J., Gippert, G. P., Reizer, J., Saier, M. H., Jr., & Wright, P. E. (1992) FEBS Lett. 296, 148-152]. 相似文献
19.
Four hundred MHz 1H-NMR and 100 MHz 13C-NMR spectra of thirteen sialylated oligosaccharide-alditols isolated from hen ovomucin and swallow nests (Collocalia mucin) were studied. The resonance assignments were determined by combining multiple-relayed coherence-transfer chemical-shift-correlated spectroscopy (multiple-Relay-Cosy) and 1H/13C chemical-shift-correlated 2-D experiments. 相似文献
20.
The 1H, 13C, and 15N backbone resonance assignments have been made for the Src homology 2 (SH2) domain of the human molecular adapter protein Grb14. The assignments, along with the majority of the non-aromatic side-chain 1H and 13C resonances are reported. The SH2 domain has been complexed with a phosphotyrosine-containing peptide (pY766) corresponding to the putative binding site in the fibroblast growth factor receptor (FGFR1). Chemical shift changes upon binding are also reported. 相似文献