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Protein Cavity (protein + cavity)
Selected AbstractsAre Sobriety and Consciousness Determined by Water in Protein Cavities?ALCOHOLISM, Issue 1 2004James R. Trudell No abstract is available for this article. [source] Specific reactions of S -nitrosothiols with cysteine hydrolases: A comparative study between dimethylargininase-1 and CTP synthetasePROTEIN SCIENCE, Issue 8 2007Oliver Braun Abstract S-Transnitrosation is an important bioregulatory process whereby NO+ equivalents are transferred between S -nitrosothiols and Cys of target proteins. This reaction proceeds through a common intermediate R,S,N(O,),S,R, and it has been proposed that products different from S -nitrosothiols may be formed in protein cavities. Recently, we have reported on the formation of such a product, an N -thiosulfoximide, at the active site of the Cys hydrolase dimethylargininase-1 (DDAH-1) upon reaction with S -nitroso- l -homocysteine (HcyNO). Here we have addressed the question of whether this novel product can also be formed with the endogenously occurring S -nitrosothiols S -nitroso- l -cysteine (CysNO) and S -nitrosoglutathione (GSNO). Further, to explore the reason responsible for the unique formation of an N -thiosulfoximide in DDAH-1 we have expanded these studies to cytidine triphosphate synthetase (CTPS), which shows a similar active site architecture. ESI-MS and activity measurements showed that the bulky GSNO does not react with both enzymes. In contrast, S-nitrosylation of the active site Cys occurred in DDAH-1 with CysNO and in CTPS with CysNO and HcyNO. Although kinetic analysis indicated that these compounds act as specific irreversible inhibitors, no N -thiosulfoximide was formed. The reasons likely responsible for the absence of the N -thiosulfoximide formation are discussed using molecular models of DDAH-1 and CTPS. In tissue extracts DDAH was inhibited only by HcyNO, with an IC50 value similar to that of the isolated protein. Biological implications of these studies for the function of both enzymes are discussed. [source] Comparison of the inward- and outward-open homology models and ligand binding of human P-glycoproteinFEBS JOURNAL, Issue 23 2009Ilza K. Pajeva An homology model of human P-glycoprotein, based on the X-ray structure of the recently resolved mouse P-glycoprotein, is presented. The model corresponds to the inward-facing conformation competent for drug binding. From the model, the residues involved in the protein-binding cavity are identified and compared with those in the outward-facing conformation of human P-glycoprotein developed previously based on the Sav1866 structure. A detailed analysis of the interactions of the cyclic peptides QZ59- RRR and QZ59- SSS is presented in both the X-ray structures of mouse P-glycoprotein and the human P-glycoprotein model generated by ligand docking. The results confirm the functional role of transmembrane domains TM4, TM6, TM10 and TM12 as entrance gates to the protein cavity, and also imply differences in their functions. The analysis of the cavities in both models suggests that the ligands remain bound to the same residues during the transition from the inward- to the outward-facing conformations. The analysis of the ligand,protein interactions in the X-ray complexes shows differences in the residues involved, as well as in the specific interactions performed by the same ligand within the same protein. This observation is supported by docking of the QZ59 ligands into human P-glycoprotein, thus aiding in the understanding of the complex behavior of P-glycoprotein substrates and inhibitors. The results confirm the possibility for multispecific drug interactions of the protein, and are important for elucidating the P-glycoprotein function and ligand interactions. [source] New insights into intracellular lipid binding proteins: The role of buried waterPROTEIN SCIENCE, Issue 10 2002Christian Lücke Abstract The crystal structures of most intracellular lipid binding proteins (LBPs) show between 5 and 20 internally bound water molecules, depending on the presence or the absence of ligand inside the protein cavity. The structural and functional significance of these waters has been discussed for several LBPs based on studies that used various biophysical techniques. The present work focuses on two very different LBPs, heart-type fatty acid binding protein (H-FABP) and ileal lipid binding protein (ILBP). Using high-resolution nuclear magnetic resonance spectroscopy, certain resonances belonging to side-chain protons that are located inside the water-filled lipid binding cavity were observed. In the case of H-FABP, the pH- and temperature-dependent behavior of selected side-chain resonances (Ser82 OgH and the imidazole ring protons of His93) indicated an unusually slow exchange with the solvent, implying that the intricate hydrogen-bonding network of amino-acid side-chains and water molecules in the protein interior is very rigid. In addition, holo H-FABP appeared to display a reversible self-aggregation at physiological pH. For ILBP, on the other hand, a more solvent-accessible protein cavity was deduced based on the pH titration behavior of its histidine residues. Comparison with data from other LBPs implies that the evolutionary specialization of LBPs for certain ligand types was not only because of mutations of residues directly involved in ligand binding but also to a refinement of the internal water scaffold. [source] Truncated hemoglobins: trimming the classical ,three-over-three' globin fold to a minimal sizeBIOCHEMISTRY AND MOLECULAR BIOLOGY EDUCATION, Issue 3 2001Mario Milani Abstract Truncated hemoglobins (trHbs) host the heme in a ,two-over-two' ,-helical sandwich which results from extensive editing of the classical ,three-over-three' globin fold. The three-dimensional structure of trHbs is based on four main ,-helices, arranged in a sort of ,-helical bundle composed of two antiparallel helix pairs (B/E and G/H). Most notably, trHbs deviate from the conventional globin fold in that they display an extended loop substituting for the heme proximal F-helix observed in globins. Moreover, since efficient adaptation of a 110,130 amino acid trHb chain to host the porphyrin ring firstly requires specific chain flexibility, trHbs contain three invariant Gly-based motifs. Inspection of the trHb three-dimensional trHb structures shows that an apparent protein cavity or tunnel would connect the protein surface to an inner region very close to the heme distal site. Such a structural feature, never observed before in (non) vertebrate globins, may have substantial implications for ligand diffusion and binding properties in trHbs. © 2001 IUBMB. Published by Elsevier Science Ltd. All rights reserved. [source] Azotobacter vinelandii Metal Storage Protein: "Classical" Inorganic Chemistry Involved in Mo/W Uptake and Release ProcessesCHEMBIOCHEM, Issue 4 2008Jörg Schemberg Dr. Abstract The release of Mo (as molybdate) from the Mo storage protein (MoSto), which is unique among all existing metalloproteins, is strongly influenced by temperature and pH value; other factors (incubation time, protein concentration, degree of purity) have minor, though significant effects. A detailed pH titration at 12,°C revealed that three different steps can be distinguished for the Mo-release process. A proportion of ,15,% at pH 6.8,7.0, an additional 25,% at pH 7.2,7.5 and ca. 50,% (up to 90,% in total) at pH 7.6,7.8. This triphasic process supports the assumption of the presence of different types of molybdenum-oxide-based clusters that exhibit different pH lability. The complete release of Mo was achieved by increasing the temperature to 30,°C and the pH value to >7.5. The Mo-release process does not require ATP; on the contrary, ATP prevents, or at least reduces the degree of metal release, depending on the concentration of the nucleotide. From this point of view, the intracellular ATP concentration is suggested to play,in addition to the pH value,an indirect but crucial role in controlling the extent of Mo release in the cell. The binding of molybdenum to the apoprotein (reconstitution process) was confirmed to be directly dependent on the presence of a nucleotide (preferably ATP) and MgCl2. Maximal reincorporation of Mo required 1 mM ATP, which could partly be replaced by GTP. When the storage protein was purified in the presence of ATP and MgCl2 (1 mM each), the final preparation contained 80 Mo atoms per protein molecule. Maximal metal loading (110,115 atoms/MoSto molecule) was only achieved, if Mo was first completely released from the native protein and subsequently (re-) bound under optimal reconstitution conditions: 1 h incubation at pH 6.5 and 12,°C in the presence of ATP, MgCl2 and excess molybdate. A corresponding tungsten-containing storage protein ("WSto") could not only be synthesized in vivo by growing cells, but could also be constructed in vitro by a metalate,ion exchange procedure by using the isolated MoSto protein. The high W content of the isolated cell-made WSto (,110 atoms/protein molecule) and the relatively low amount of tungstate that was released from the protein under optimal "release conditions", demonstrates that the W-oxide-based clusters are more stable inside the protein cavity than the Mo-oxide analogues, as expected from the corresponding findings in polyoxometalate chemistry. The optimized isolation of the W-loaded protein form allowed us to get single crystals, and to determine the crystal X-ray structure. This proved that the protein contains remarkably different types of polyoxotungstates, the formation of which is templated in an unprecedented process by the different protein pockets. (Angew. Chem. Int. Ed.2007, 46, 2408,2413). [source] | ||||||||||