1,721,208 research outputs found
C-13 AND H-1-NMR STUDIES OF IMIDAZOLE BINDING TO NATIVE AND CO(II)-SUBSTITUTED HUMAN CARBONIC ANHYDRASE-I
No full textThe inhibition constant of imidazole toward human carbonic anhydrase I has been directly determined at pH 6.8 and 8.8 through C-13 NMR. The data are in agreement with those obtained by indirect methods and confirm that the binding affinity of imidazole is substantially constant in the pH range investigated. Analysis of the isotropically-shifted signals in the H-1 NMR spectra of the Co(II)-substituted enzyme interacting with imidazole at high and low pH in the presence of sulfate as counterion indicates the likely formation of a five-coordinate HCAI-imidazole-sulfate adduct at low pH. The binding mode of imidazole in the active site of the enzyme is discussed
Il giardino e le mura. Ai confini fra natura e storia. Atti del Convegno di studi (S. Miniato Alto, 23-24 giugno 1995)
No full textA structural and dynamic characterization of the EF-hand protein CLSP
No full textHuman Calmodulin-Like Skin Protein (CLSP) is a 15.92 kDa protein expressed during the differentiation of keratinocytes to corneocytes. The structure and dynamics of this protein have been characterized by NMR spectroscopy. CLSP is affected by high mobility both in the fast (ps-ns) and intermediate (μs) timescale range, with the N-terminal and C-terminal domains being differently affected. The isolated N-terminal and C-terminal domains were also expressed and analyzed. The structure of the isolated C-terminal domain is presented, which is very similar to that in the entire protein. The N-terminal domain is characterized by four stable helices which experience large fluctuations. This is shown to be due to mutations in the hydrophobic core. Fast mobility is experienced in the two loops. Almost no long range NOEs are observed. The overall N-terminal domain behavior is similar both in the full length protein and in the isolated domain. By exploiting the capability of Tb3+ bound to CLSP to induce partial orientation of the molecule in a magnetic field, restricted motion of one domain with respect to the other was proved. By using NMR, ITC and ESI-MS the calcium and magnesium binding properties were investigated. Finally CLSP is framed into the evolutionary scheme of the Calmodulin-like famil
INFLUENCE OF SURFACE-CHARGES ON REDOX PROPERTIES IN HIGH-POTENTIAL IRON-SULFUR PROTEINS
No full textThe pH-dependence of the reduction potential determined through differential pulse voltammetry for the high potential iron sulfur proteins (HiPIP) from R. globiformis, C. vinosum, R. gelatinosus, E. vacuolata (I and II), E. halophila (I and II) is reported. A decrease in reduction potential with pH is invariably observed in the pH range where deprotonation of the imidazolium nitrogen of histidine residue(s) occurs. No pH dependence is observed for the only protein lacking histidines. It appears that surface charges like the His imidazolium groups are capable of influencing the reduction potential despite the known quencing of the electrostatic interactions due to solvent effects
POLYMETALLIC HYDROLYTIC ZINC ENZYMES - PROBING THE SITE OF NUCLEASE P1 THROUGH COBALT(II) SUBSTITUTION
No full textPartial Co(II) substitution in the three-zinc site of P. citrinum nuclease P1 has been achieved. The Co(II) ion is found to bind to the site of the most EDTA-labile Zn atom with an 80% site occupancy. An affinity constant of 10(5) M(-1) for metal binding was determined from the visible spectra which also indicate a six-coordinate Co(II) geometry. The hyperfine-shifted H-1 NMR resonances suggest that metal substitution occurred at the Zn3 site (X-ray atom numbering)
H-1-NMR SPECTROSCOPY AND THE ELECTRONIC-STRUCTURE OF THE HIGH-POTENTIAL IRON SULFUR PROTEIN FROM CHROMATIUM-VINOSUM
No full textH-1 NMR and NOE measurements have been performed on reduced HiPIP from Chromatium vinosum. 1D and 2D saturation transfer experiments have allowed a full correlation between the isotropically shifted signals of the reduced and oxidized species. The pairwise assignment of the cysteine geminal beta-CH2 protons is performed. This allows the relative evaluation of the expectation value [S(iz)] for each iron ion i as well as its temperature dependence. A theoretical approach has been able to account for upfield shifts of beta-CH2 cysteine protons in the oxidized species. The H-1 NMR data turn out to be a valuable benchmark for testing theoretical models for Fe4S4 clusters. The present model supports the existence of a mixed-valence pair with S = 9/2 ground state in the oxidized protein and is consistent with the presence of analogous mixed-valence pairs also in the reduced protein. The NMR data on the oxidized protein also show that, even at room temperature, electron delocalization mainly occurs within one particular Fe(II)-Fe(III) pair
NMR for sample quality assessment in metabolomics
No full textThe EU Framework 7 project SPIDIA was the occasion for development of NMR approaches to evaluate the impact of different pre-analytical treatments on the quality of biological samples dedicated to metabolomics. Systematic simulation of different pre-analytical procedures was performed on urine and blood serum and plasma. Here we review the key aspects of these studies that have led to the development of CEN technical specifications, to be translated into ISO/IS in the course of the EU Horizon 2020 project SPIDIA4P. Inspired by the SPIDIA results, follow-up research was performed, extending the analysis to different sample types and to the different effects of long-term storage. The latter activity was in conjunction with the local European da Vinci Biobank. These results (which partially contributed to the ANNEX of CEN/TS 16945“MOLECULAR IN VITRO DIAGNOSTIC EXAMINATIONS - SPECIFICATIONS FOR PRE-EXAMINATION PROCESSES FOR METABOLOMICS IN URINE, VENOUS BLOOD SERUM AND PLASMA”)are presented in detail
Solvent 1H NMRD study of biotinylated paramagnetic liposomes containing Gd-bis-SDA-DTPA or Gd-DMPE-DTPA
No full textMAGNETIC-RESONANCE OF FE-S CLUSTERS - ISOLATION AND CHARACTERIZATION OF A 7FE FERREDOXIN FROM RHODOPSEUDOMONAS-PALUSTRIS
No full textA novel iron-sulfur protein from the photosynthetic purple bacterium Rhodopseudomonas palustris was purified to homogeneity and identified as a ferredoxin on the basis of its physicochemical properties. Based on the uv/vis spectrum, iron quantitation, cyclic voltammetry, EPR, and H-1 NMR data, the ferredoxin is found to contain two iron-sulfur clusters, one [3Fe-4S] and one [4Fe-4S], which places this protein in the class of 7Fe ferredoxins. The voltammetric peak potentials of the two clusters are -0.260 and -0.560 V at pH 8.0. The molecular mass around 19 kDa makes this protein the heaviest known in this class. This paper further demonstrates the diagnostic power of magnetic resonance spectroscopies in recognition of the two types of clusters in iron-sulfur proteins
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