Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The structure of the Neurospora crassa plasma membrane H(+)-ATPase has been investigated using a variety of chemical and physiochemical techniques. The transmembrane topography of the H(+)-ATPase has been elucidated by a direct, protein chemical approach. Reconstituted proteoliposomes containing purified H(+)-ATPase molecules oriented predominantly with their cytoplasmic surface facing outward were treated with trypsin, and the numerous peptides released were purified by HPLC and subjected to amino acid sequence analysis. In this way, seventeen released peptides were unequivocally identified as located on the cytoplasmic side of the membrane, and numerous intervening segments could be inferred to be cytoplasmically located by virtue of the fact that they are too short to cross the membrane and return between sequences established to be cytoplasmically located. Additionally, three large membrane-embedded segments of the H(+)-ATPase were isolated using our recently developed methods for purifying hydrophobic peptides, and identified by amino acid sequence analysis. This information established the topographical location of virtually all of the 919 residues in the H(+)-ATPase molecule, allowing the formulation of a reasonably detailed model for the transmembrane topography of the H(+)-ATPase polypeptide chain. Separate studies of the cysteine chemistry of the H(+)-ATPase have demonstrated the existence of a single disulfide bridge in the molecule, linking the NH2- and COOH-terminal membrane-embedded domains. And, analyses of the circular dichroism and infrared spectra of the purified H(+)-ATPase have elucidated the secondary structure composition of the molecule. A first-generation model for the tertiary structure of the H(+)-ATPase based on this information and other considerations is presented.
Mol Cell Biochem 1992 Sep 08
PMID:Probing the structure of the Neurospora crassa plasma membrane H(+)-ATPase. 146 Dec 58

A number of ligands for the selective purification by affinity chromatography of the trypsin-like protease, porcine pancreatic kallikrein, were designed de novo by computer-aided molecular design. The ligands were designed to mimic the side-chains of a number of arginyl dipeptides and included a benzamidine moiety substituted on a triazine ring. The ligands displayed inhibitory activities against pancreatic kallikrein which mirrored the specificity constants of the dipeptides they were designed to mimic. The ligand with the highest affinity for the enzyme, an analogue of a Phe-Arg dipeptide, when immobilized to Sepharose CL-4B via a hexamethylene spacer arm, purified pancreatic kallikrein 110-fold in one step from a crude pancreatic acetone extract.
J Mol Recognit 1992 Jun
PMID:Design of novel affinity adsorbents for the purification of trypsin-like proteases. 147 81

The 21 kDa protein of liver from Atlantic salmon (Salmo salar) has been purified. Hepatic nuclei were extracted with 0.75 M HClO4. The extracted proteins were fractionated using reversed phase high performance liquid chromatography. The purity of the protein was analysed by isoelectric focusing in the first, and SDS-polyacrylamide gel electrophoresis in the 2nd dimension. Isoelectric focusing separated the protein into 5 spots. In gel trypsin digestion after isoelectric focusing followed by SDS-polyacrylamide gel electrophoresis resulted in identical migration of the tryptic peptides. The amino acid composition of the 21 kDa protein was similar to that of high mobility group (HMG) proteins C and D from rainbow trout (Oncorhynchus mykiss). The N-terminal sequence of the amino acids 1-19 revealed a conserved region characteristic for HMG 14/17 proteins of mammals and avians, and their equivalents in rainbow trout. Considering the electrophoretic mobility, amino acid composition and N-terminal amino acid sequence it is concluded that the 21 kDa protein of Atlantic salmon is a member of the HMG protein family resembling the HMG D protein of rainbow trout.
Cell Mol Biol (Noisy-le-grand) 1992 Nov
PMID:Purification of the Atlantic salmon hepatic 21 kDa protein and classification as a high mobility group chromatin protein. 147 4

The study presented herein is a bidimensional approach to the complementarity of two molecular surfaces. From two chosen sections we have established a methodology of generating the optimal matching of two shapes. Our approach consists in describing two molecular surface sections by a shape vector (the angular profile), in finding their matching patterns by comparison of the two profiles, and in optimizing the relative locations of the two sections in two-dimensional space, using rotations and translations defined by geometric characteristics. The set of optimal configurations are successively displayed on a screen. Satisfying results have been obtained for the matching of the complex kallikreine A-trypsin pancreatic bovin 2. This efficient method could be used as a preprocessing for a tridimensional shape complementarity approach between two molecular surfaces.
J Mol Graph 1992 Dec
PMID:Searching for geometric molecular shape complementarity using bidimensional surface profiles. 147 92

Hexokinase II prepared from Ehrlich-Lettre hyperdiploid tumor cells (ELD cells) was subjected to a limited digestion by trypsin. After 60 min digestion, hexokinase II (100 kDa) was completely cleaved to two fragments with the molecular weight of about 60 kDa and 40 kDa as manifested in SDS-PAGE. It was noteworthy that the enzyme activity was observed even at the time when the native enzyme molecule was no more detectable. These fragments were separated by SDS-PAGE irrespective of the presence of a reducing agent, but neither by native PAGE nor by cellulose acetate membrane electrophoresis under the nondenaturing conditions. Neither kinetic parameters such as Km values for ATP and glucose nor an ability of binding to mitochondria were changed significantly by the tryptic digestion. These results indicate that an essential conformation of hexokinase II can be restored by the self-association of two fragments produced as a result of the cleavage by trypsin at the middle of the molecule. Affinity labeling with 2',3'-dialdehyde ATP followed by the trypsin digestion showed that ATP binding site resided in the 40 kDa fragment. Furthermore, the mode of the response in the incorporation of this ATP analog to hexose phosphate, moreover, was similar to that in the catalytic activity.
Mol Cell Biochem 1992 Nov 04
PMID:Cleavage of hexokinase II to two domains by trypsin without significant change in catalytic activity. 148 Jan 68

The bovine adrenal cortex adrenodoxin gene was inserted into pTZ19 under T7 promoter control. The adrenodoxin mRNA was synthesized with T7 RNA polymerase and then translated in the reticulocyte cell-free translation system. The protein product was identified as the adrenodoxin precursor with molecular weight 22000. The import of the precursor into isolated yeast mitochondria was carried out. The protein was found to be inserted into the trypsin-insensitive compartment of mitochondria via an energy dependent way. This resulted in the processing of the precursor to the 12000-mature form. Thus, the precursor of mammalian adrenodoxin can be normally imported into yeast mitochondria.
Mol Biol (Mosk)
PMID:[Synthesis of a bovine adrenodoxin precursor in vitro and its import into yeast mitochondria]. 150 62

Previous studies have demonstrated that protease treatment of zona-free mouse eggs impairs sperm-egg interaction (Boldt et al.: Biol Reprod 39:19-27, 1988) and causes modification of a 94 kD egg plasma membrane protein (Boldt et al., Gamete Res 23:91-101, 1989). In this report, the ability of eggs to recover penetration ability following protease treatment was examined. Zona-free mouse eggs were isolated and treated with either trypsin or chymotrypsin (1 mg/ml, 20 min), then cultured for 0, 3, or 6 hr before insemination. Eggs cultured for 3 or 6 hr displayed significantly higher penetration levels than eggs inseminated immediately after protease treatment, indicating a recovery of penetration ability during the 3 or 6 hr incubation period. The recovery of penetration ability was not blocked by inclusion of cyclohexamide (50 micrograms/ml) during the 3 or 6 hr culture period, indicating that protein synthesis was not required for recovery of fusion ability. Cell surface radiolabeling studies with 125I revealed that a 94 kD cell surface protein was lost immediately following trypsin or chymotrypsin treatment but was found on the egg surface after the 3 or 6 hr recovery period. Recovery of the 94 kD egg surface protein occurred in the presence of cyclohexamide, and metabolic radiolabeling studies with 35S-methionine confirmed that synthesis of a 94 kD protein was blocked by cyclohexamide. These results suggest that the recovery of penetration ability after protease treatment of zona-free eggs is due to recovery of the 94 kD cell surface protein, providing further evidence for the involvement of the 94 kD protein in sperm-egg interaction.
Mol Reprod Dev 1992 Sep
PMID:Recovery of penetration ability in protease-treated zona-free mouse eggs occurs coincident with recovery of a cell surface 94 kD protein. 151 Aug 43

Well-diffracting crystals of bovine epsilon-thrombin in complex with several "non-peptidic" benzamidine and arginine-based thrombin inhibitors have been obtained by co-crystallization. The 2.3 A crystal structures of three complexes formed either with NAPAP, 4-TAPAP, or MQPA, were solved by Patterson search methods and refined to crystallographic R-values of 0.167 to 0.178. The active-site environment of thrombin is only slightly affected by binding of the different inhibitors; in particular, the exposed "60-insertion loop" essentially maintains its typical projecting structure. The D-stereoisomer of NAPAP and the L-stereoisomer of MQPA bind to thrombin with very similar conformations, as previously inferred from their binding to bovine trypsin; the arginine side-chain of the latter inserts into the specificity pocket in a "non-canonical" manner. The L-stereoisomer of 4-TAPAP, whose binding geometry towards trypsin was only poorly defined, is bound to the thrombin active-site in a compact conformation. In contrast to NAPAP, the distal p-amidino/guanidino groups of 4-TAPAP and MQPA do not interact with the carboxylate group of Asp189 in the thrombin specificity pocket in a "symmetrical" twin N-twin O manner, but through "lateral" single N-twin O contacts; in contrast to the p-amidino group of 4-TAPAP, however, the guanidyl group of MQPA packs favourably in the pocket due to an elaborate hydrogen bond network, which includes two entrapped water molecules. These thrombin structures confirm previous conclusions of the important role of the intermolecular hydrogen bonds formed with Gly216, and of the good sterical fit of the terminal bulky hydrophobic inhibitor groups with the hydrophobic aryl binding site and the S2-cavity, respectively, for tight thrombin active site binding of these non-peptidic inhibitors. These accurate crystal structures are presumed to be excellent starting points for the design and the elaboration of improved antithrombotics.
J Mol Biol 1992 Aug 20
PMID:Refined 2.3 A X-ray crystal structure of bovine thrombin complexes formed with the benzamidine and arginine-based thrombin inhibitors NAPAP, 4-TAPAP and MQPA. A starting point for improving antithrombotics. 151 46

The crystal structure of trypsin-modified elongation factor Tu from Escherichia coli, in complex with the cofactor guanosine diphosphate has been refined to a crystallographic R-factor of 19.3%, at 2.6 A resolution. In the model described, the root-mean-square deviation from ideality is 0.019 A for bond distances and 3.9 degrees for angles. The protein consists of three domains: an alpha/beta domain (residues 1 to 200), containing the binding site of the GDP cofactor, and consisting of a six-stranded beta-pleated sheet, six alpha-helices, and two all-beta domains (residues 209 to 299 and 300 to 393), belonging to the tertiary structural class of antiparallel beta-barrels. The GDP-binding domain has a folding that is found in other GDP-binding proteins. Elongation factor Tu interacts with proteins, nucleic acids and nucleotides, making this molecule well suited as a model system for the study of these interactions.
J Mol Biol 1992 Feb 05
PMID:Refined structure of elongation factor EF-Tu from Escherichia coli. 154 16

A procedure for separation of the catalytic and regulatory subunits of sterol sulphate sulphohydrolase from human placenta microsomes with the use of Concanavalin A-Sepharose chromatography is presented. The Km value for the catalytic subunit with oestrone sulphate is 1.2 x 10(-5) M. The Hill coefficient value h, for the reconstituted enzyme complex is 3, the S0.5 = 0.68 x 10(-3) M and the value of Km is 0.31 x 10(-12) M. The regulatory subunit is trypsin sensitive, while the catalytic one is resistant to trypsin digestion.
J Steroid Biochem Mol Biol 1992 Feb
PMID:Subunits of sterol sulphate sulphohydrolase from human placenta microsomes. 154 81


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