SEARCH · PubMed Health

Results for “BINDING SITES”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 73 records · Page 4Linked to original sources

PubMed1988

Solubilization and characterization of substance P-binding sites from chick brain membranes.

Sites binding monoiodinated-Bolton-Hunter-reagent-labelled substance P were solubilized from 1-day-old-chick brain membrane by using non-ionic detergents (1% digitonin/1% n-octyl glucoside) and a high concentration of NaCl (0.5 M). The solubilized preparation retained the pharmacological properties of the high-affinity binding sites found in the native membrane. The high density of specific binding sites (approximately 2 pmol of binding sites/mg of protein) suggests that the chick brain membranes may be a useful source for the purification of the substance P-binding sites.

Adenosine Triphosphate↗

PubMed1990

Resonance energy transfer study on the proximity relationship between the GTP binding site and the rifampicin binding site of Escherichia coli RNA polymerase.

Terbium (III) upon complexation with guanosine 5'-triphosphate showed remarkable enhancement of fluorescence emission at 488 and 545 nm when excited at 295 nm. Analysis of the binding data yielded a value for the mean Kd between Tb(III) and GTP of 0.2 microM, with three binding sites for Tb(III) on GTP. 31P and 1H NMR measurements revealed that Tb(III) mainly binds the phosphate moiety of GTP. Fluorescence titration of the emission signals of the TbGTP complex with varying concentrations of Escherichia coli RNA polymerase resulted in a Kd value of 4 microM between the TbGTP and the enzyme. It was observed that TbGTP can be incorporated in the place of GTP during E. coli RNA polymerase catalyzed abortive synthesis of dinucleotide tetraphosphate at T7A2 promoter. Both the substrate TbGTP and the inhibitor of the initiation of transcription rifampicin bind to the beta-subunit of E. coli RNA polymerase. This allows the measurement of the fluorescence excited-state energy transfer from the donor TbGTP-RNA polymerase to the acceptor rifampicin. Both emission bands of Tb(III) overlap with the rifampicin absorption, and the distances at 50% efficiency of energy transfer were calculated to be 28 and 24 A for the 488- and 545-nm emission bands, respectively. The distance between the substrate binding site and the rifampicin binding site on the beta-subunit of E. coli RNA polymerase was measured to be around 30 A. This suggests that the nature of inhibition of transcription by rifampicin is essentially noncompetitive with the substrate.

Binding Sites↗

PubMed1989

The Fc binding site for streptococcal protein G is in the C gamma 2-C gamma 3 interface region of IgG and is related to the sites that bind staphylococcal protein A and human rheumatoid factors.

The isolated 35 kDa fragment of protein G obtained by papain digestion of group G streptococci was found to bind solid phase intact IgG, Fc (2C gamma 2 + 2C gamma 3 domains), F(ab')2 and F(acb)2 (F(ab')2 + 2C gamma 2 domains) fragments but not pFc' (2C gamma 3 domains) fragments. The level of binding to rabbit F(acb)2 and rabbit F(ab')2 fragments was similar. Protein G binding to solid phase Fc fragments was inhibited by IgG, Fc, staphylococcal protein A and its monovalent fragment D, but was enhanced by F(ab')2 fragments. Chemical modification of tyrosine but not histidine residues of IgG abrogated its ability to inhibit the binding of protein G to solid phase Fc fragments. Protein G was found to strongly inhibit the binding of a monoclonal and a polyclonal human rheumatoid factor to IgG. These findings indicate that protein G binds with separate sites to the Fc and F(ab')2 fragments of IgG, that the interaction with the Fc fragment occurs at the C gamma 2-C gamma 3 domain interface region and that tyrosine but not histidine residues in this area are likely involved. The relationship of the Fc fragment-binding site specificity of protein G to that of other microbial IgG binding proteins and human rheumatoid factors is discussed.

Animals↗

PubMed2006

Allosteric interactions between the nucleotide-binding sites and the ssDNA-binding site in the PriA helicase-ssDNA complex. 3.

Allosteric interactions between the strong and weak nucleotide-binding sites and the total and proper single-stranded (ss)DNA-binding sites of the Escherichia coli PriA helicase have been analyzed using the fluorescence titration technique. Binding of the DNA exclusively to the proper DNA-binding site of the helicase, profoundly affects the intrinsic affinities of both nucleotide-binding sites, indicating a direct communication between the nucleotide-binding sites and the proper DNA-binding site. The communication involves conformational changes of the entire protein molecule. Nevertheless, the bound DNA differently affects the structures of the strong and weak nucleotide-binding sites. While the polarity of the strong site is moderately diminished, the polarity of the weak site is dramatically increased, indicating an intimate involvement of the weak site in controlling the helicase interactions with the DNA. The strong site does not directly control the DNA affinity of the enzyme. Only when the helicase has both nucleotide-binding sites saturated with ADP but not with ATP analogues does the enzyme have an increased affinity for the ssDNA, indicating that the control of ssDNA affinity involves a coordinated action of both nucleotide-binding sites and depends upon the phosphate group of the bound cofactor. A dramatic increase of the DNA affinity, when the DNA encompasses the total DNA-binding site of the enzyme, with both nucleotide-binding sites saturated with ADP or NDP, indicates that an additional area of the protein within the total DNA-binding site becomes engaged in interactions with the DNA. The significance of these results for the enzyme activities in the DNA unwinding and recognition is discussed.

Adenosine Diphosphate↗

PubMed1998

Anatomy of lipase binding sites: the scissile fatty acid binding site.

Shape and physico-chemical properties of the scissile fatty acid binding sites of six lipases and two serine esterases were analyzed and compared in order to understand the molecular basis of substrate specificity. All eight serine esterases and lipases have similar architecture and catalytic mechanism of ester hydrolysis, but different substrate specificities for the acyl moiety. Lipases and esterases differ in the geometry of their binding sites, lipases have a large, hydrophobic scissile fatty acid binding site, esterases like acetylcholinesterase and bromoperoxidase have a small acyl binding pocket, which fits exactly to their favorite substrates. The lipases were subdivided into three sub-groups: (1) lipases with a hydrophobic, crevice-like binding site located near the protein surface (lipases from Rhizomucor and Rhizopus); (2) lipases with a funnel-like binding site (lipases from Candida antarctica, Pseudomonas and mammalian pancreas and cutinase); and (3) lipases with a tunnel-like binding site (lipase from Candida rugosa). The length of the scissile fatty acid binding site varies considerably among the lipases between 7.8 A in cutinase and 22 A in Candida rugosa and Rhizomucor miehei lipase. Location and properties of the scissile fatty acid binding sites of all lipases of known structure were characterized. Our model also identifies the residues which mediate chain length specificity and thus may guide protein engineering of lipases for changed chain length specificity. The model was supported by published experimental data on the chain length specificity profile of various lipases and on mutants of fungal lipases with changed fatty acid chain length specificity.

Animals↗

PubMed2003

Identification of HTF (HER2 transcription factor) as an AP-2 (activator protein-2) transcription factor and contribution of the HTF binding site to ERBB2 gene overexpression.

The ERBB2 gene is overexpressed in 30% of human breast cancers and this is correlated with poor prognosis. Overexpression of the ERBB2 gene is due to increased transcription and gene amplification. Our previous studies have identified a new cis element in the ERBB2 promoter which is involved in the gene's overexpression. This cis element, located 501 bp upstream from the main ERBB2 transcription initiation site, binds a transcription factor called HTF (HER2 transcription factor). We report here the identification of HTF as an AP-2 (activator protein-2) transcription factor. The new cis element is bound by AP-2 with high affinity, compared with a previously described AP-2 binding site located 284 bp downstream. Co-transfection of an AP-2alpha expression vector with a reporter vector containing the newly identified AP-2 binding site in front of a minimal ERBB2 promoter induced a dose-dependent increase in transcriptional activity. We examined the contribution of the new AP-2 binding site to ERBB2 overexpression. For this purpose we abolished the new and/or the previously described AP-2 binding sequence by site-directed mutagenesis. The results show that the two functional AP-2 sites in the first 700 bp of the ERBB2 promoter co-operate to achieve maximal transcriptional activity.

Base Sequence↗

PubMed1983

Comparison of ribosomal entry and acceptor transfer ribonucleic acid binding sites on Escherichia coli 70S ribosomes. Fluorescence energy transfer measurements from Phe-tRNAPhe to the 3' end of 16S ribonucleic acid.

Distances were measured by nonradiative energy transfer from fluorescent probes specifically located on one of three points of yeast or Escherichia coli Phe-tRNAPhe enzymatically bound to the entry site or to the acceptor site of E. coli 70S ribosomes to energy-accepting probes on the 3' end of the 16S ribonucleic acid (RNA) of the 30S subunit. The Y base in the anticodon loop of yeast tRNAPhe was replaced by proflavin. Fluorescein isothiocyanate was attached to the X base (position 47) of E. coli tRNAPhe. E. coli tRNAPhe which had been photochemically cross-linked between positions 8 and 13 followed by chemical reduction to form a fluorescent probe was also used. Labeled tRNAs were aminoacylated and enzymatically bound to the ribosome in the presence of elongation factor Tu and guanosine 5'-triphosphate (acceptor-site binding) or a nonhydrolyzable analogue (entry-site binding). Nonradiative energy transfer measurements were made of the distances between fluorophores located on the Phe-tRNA and the fluorophore at the 3' end of 16S RNA. Calculations were based on comparison of the fluorescence lifetime of the energy donor, located on the Phe-tRNA, in the absence and presence of an energy acceptor on the 3' end of the 16S RNA. Under both sets of binding conditions, the distances to the 3' end of 16S RNA were found to be the following: cross-linked tRNA, greater than 69 A; Y base of tRNA, greater than 61 A. The distance between the 3' end of 16S RNA and the X base of tRNA was found to be 81 A under acceptor-site binding conditions but greater than 86 A under entry-site binding conditions.

Energy Transfer↗

PubMed1993

Conversion of a magnesium binding site into a zinc binding site by a single amino acid substitution in Escherichia coli alkaline phosphatase.

The replacement of aspartic acid by histidine at position 153 in Escherichia coli alkaline phosphatase results in a mutant enzyme that is remarkably similar to certain mammalian alkaline phosphatases that are activated by magnesium in a time-dependent fashion. These mammalian alkaline phosphatases have histidine at the position corresponding to 153 of the E. coli sequence. Here we report the three-dimensional structure of the mutant E. coli alkaline phosphatase with histidine at position 153. The structure reveals that the octahedral magnesium binding site has been converted to a tetrahedral zinc binding site with an imidazole ring nitrogen of His-153 as one of the ligands to the zinc. The alteration in metal binding caused by the mutation could explain the origin of the magnesium activation observed with the mammalian alkaline phosphatases. The structure also reveals differences in the mode of phosphate binding, explaining the enhanced phosphate affinity and the reduced activity of the mutant enzyme in the presence of zinc.

Alkaline Phosphatase↗

PubMed1990

Serotonin binding sites. II. Muramyl dipeptide binds to serotonin binding sites on myelin basic protein, LHRH, and MSH-ACTH 4-10.

Previously, we reported the existence of structurally similar serotonin binding sites on myelin basic protein, LHRH, and MSH-ACTH 4-10. We now report that the adjuvant peptide, muramyl dipeptide (N-acetyl-muramyl-L-Ala-D-isoGln) also binds to these sites. This observation may help to explain previous observations of serotonin-like activity by muramyl peptides, including the promotion of slow-wave sleep and fever induction. The observation may also provide an important link between the immune system and the nervous system that may explain the role of muramyl dipeptide adjuvants in causing autoimmune diseases to serotonin-regulated proteins and their receptors, as well as the alterations in serotonin levels that are often observed in autoimmune diseases. The observation provides concrete evidence for a dual-antigen hypothesis for the induction of autoimmune diseases by an adjuvant-peptide complex. Application of such a mechanism for induction of autoimmunity may be of importance in understanding a number of postinfectious and postvaccinal neuropathies, and suggests a possible etiology for autism, in which many patients have high blood serotonin levels, autoimmune reactions to myelin basic protein, and antibodies to serotonin binding sites. Finally, the observation suggests that glycopeptides may act as neurotransmitters.

Acetylmuramyl-Alanyl-Isoglutamine↗

PubMed1992

Low-affinity Ca(2+)-binding sites versus Zn(2+)-binding sites in histidine-rich Ca(2+)-binding protein of skeletal muscle sarcoplasmic reticulum.

Histidine-rich Ca(2+)-binding protein (HRC) is a 170 kDa protein that can be identified in the isolated sarcoplasmic reticulum from rabbit skeletal muscle by its ability to bind [125I]low-density lipoprotein on blots after SDS-PAGE and that appears to be bound to the junctional membrane through calcium bridges. Molecular cDNA cloning of this protein predicts the existence of a Ca(2+)-binding domain and of a distinct heavy-metal binding domain at the cystein-rich COOH-terminus. Here we demonstrate, using radioactive ligand blot techniques, that HRC protein binds 45Ca at low affinity, as well as being able to bind 65Zn, but at different sites, that are largely inhibitable by prior reductive alkylation of the protein. In contrast to Ca(2+)-binding protein calsequestrin not having detectable 65Zn-binding sites, HRC protein bound selectively to immobilized Zn2+ on IDA-agarose affinity columns. Our results also indicate that rabbit and human 140 kDa HRC protein have common properties.

Animals↗

PubMed1991

In vitro autoradiographic endothelin-1 binding sites and sarafotoxin S6B binding sites in rat tissues.

1. The distribution of binding sites for [125I]-labelled endothelin-1 ([125I]-ET-1) and [125I]-labelled sarafotoxin S6B ([125I]-SRT) was visualized in rat tissues using in vitro autoradiography. 2. A high density of endothelin-1 (ET-1) binding was found in the heart. In the kidney, ET-1 binding occurred in association with glomeruli, proximal tubules, the inner stripe and inner medulla. In the adrenal, a high density of ET-1 binding occurred in the medulla as well as the zona glomerulosa. 3. The binding affinity constant (KA) for ET-1 binding in these sites ranged from 1 to 10 x 10(9)/mol per litre. 4. Although sarafotoxin S6B (SRT) was 10-100-fold weaker than ET-1 in displacing [125I]-ET-1 from these sites, 1 mumol/L unlabelled SRT completely abolished [125I]-ET-1 binding in all sites. Other venom peptides did not affect [125I]-ET-1 binding. 5. The pattern of [125I]-SRT receptor binding in rat tissues by in vitro autoradiography was identical to that for ET-1 receptor binding, and both unlabelled SRT and unlabelled ET-1 fully competed with [125I]-SRT for binding. 6. These results provide evidence that SRT binds to the ET receptor in a range of rat tissues. The results suggest that there may be subclasses of ET receptors which can be distinguished by the relative potencies of ET-1 and SRT at various tissues.

Adrenal Glands↗

PubMed2002

A neutral molecule in a cation-binding site: specific binding of a PEG-SH to acetylcholinesterase from Torpedo californica.

The crystal structure of acetylcholinesterase from Torpedo californica complexed with the uncharged inhibitor, PEG-SH-350 (containing mainly heptameric polyethylene glycol with a terminal thiol group) is determined at 2.3 A resolution. This is an untypical acetylcholinesterase inhibitor, since it lacks the cationic moiety typical of the substrate (acetylcholine). In the crystal structure, the elongated ligand extends along the whole of the deep and narrow active-site gorge, with the terminal thiol group bound near the bottom, close to the catalytic site. Unexpectedly, the cation-binding site (formed by the faces of aromatic side-chains) is occupied by CH(2) groups of the inhibitor, which are engaged in C-H...pi interactions that structurally mimic the cation-pi interactions made by the choline moiety of acetylcholine. In addition, the PEG-SH molecule makes numerous other weak but specific interactions of the C-H...O and C-H...pi types.

Acetylcholinesterase↗

PubMed1984

Nd3+ and Co2+ binding to sarcoplasmic reticulum CaATPase. An estimation of the distance from the ATP binding site to the high-affinity calcium binding sites.

Nd3+ binding to sarcoplasmic reticulum (SR) was detected by inhibition of ATPase activity and directly by a fluorimetric assay. Both methods indicated that Nd3+ inhibited the ATPase activity by binding in the high-affinity Ca2+ binding sites. The stoichiometry of binding was about 11 nmol of Nd3+ bound per mg of SR proteins at pNd = 6.5. At higher [Nd3+], substantial nonspecific binding occurred. The association constant for Nd3+ binding to the high-affinity Ca2+ binding sites was estimated to be near 2 X 10(9) M-1. When the CaATPase was inactivated with fluorescein isothiocyanate (FITC), 5.3 nmol were bound per mg of SR protein. This fluorescent probe is known to bind in the ATP binding site. The stoichiometry of Nd3+ binding to FITC-labeled CaATPase was the same, within experimental error, as to the unlabeled CaATPase. Fluorescence energy transfer between FITC in the ATP site and Nd3+ in the Ca2+ sites was found to be very small. This donor-acceptor pair has a critical distance of 0.93 nm and the distance between the ATP site and the closest Ca2+ was estimated to be greater than 2.1 nm. Parallel measurements with FITC-labeled SR and Co2+, an acceptor with a critical distance 1.2 nm, suggested the ATP and Ca2+ binding sites are greater than 2.6 nm apart.

Animals↗

PubMed1989

Brain leukotriene C4 binding sites are S-alkylglutathione binding sites.

Leukotriene C4 binding to mouse brain membranes was readily displaced by S-alkylglutathione derivatives, with the affinity of the test compound increasing as the alkyl chain length increases. S-decylglutathione was as potent as leukotriene C4. These data suggest that brain membrane leukotriene C4 binding sites are S-alkylglutathione binding sites, possibly microsomal glutathione transferase.

Alkylation↗

PubMed1993

Studies on the mode of antifungal action of pradimicin antibiotics. II. D-mannopyranoside-binding site and calcium-binding site.

Based on the structure-activity relationship data of BMY-28864 and related pradimicin derivatives, the calcium salt-forming ability and the D-mannopyranoside-specific visible absorption maximum shift of BMY-28864 were analysed in the ternary complex formation of BMY-28864 with D-mannopyranoside and calcium. The free C-18 carboxyl group of BMY-28864 was proved to be the sole site for binding to calcium, while no hydroxyl groups of the aglycone were involved in calcium salt formation. The stereospecific D-mannopyranoside-recognizing ability of BMY-28864 was completely abolished by removal of the C-5 disaccharide moiety, and, more particularly, of the C-5 thomosamine moiety. Close relationship of these findings with the antifungal action was also supported by the in vitro antifungal assay and the potassium leakage induction test.

Anthracyclines↗

PubMed1987

The membrane of the rough endoplasmic reticulum contains cytoplasmically exposed high affinity GTP-binding sites.

Binding experiments using [14C]GTP and rat liver rough microsomes gave Scatchard plots with Kd congruent to 0.1 microM and a binding capacity congruent to 40 pmol/mg microsomal protein. Removal of the ribosomes did not modify the binding parameters. GTP was competed out by GTP analogues but not by ATP. Limited proteolysis of rough microsomes decreased the GTP-binding capacity and prevented GTP from suppressing the latency of mannose-6-phosphatase and of the synthesis of dolichol-linked chitobiose. The GTP-binding protein is probably involved in these effects of GTP. Its function could be to act in the association-dissociation of membrane components at the ribosome-membrane junction.

Animals↗

PubMed1981

The maturation of cortical serotonin binding sites.

Binding of serotonin to rat brain membranes increased linearly from birth to adulthood, but newborn receptor densities were already 39% of adult levels. These data suggest a postnatal development of serotonin receptors, coincident with synaptic maturation but do not preclude a non-transmitter function for serotonin during early maturation.

Aging↗

PubMed1991

Mutating protein kinase cAMP-binding sites into cGMP-binding sites. Mechanism of cGMP selectivity.

The cAMP-dependent protein kinase contains two different cAMP-binding sites referred to as the slow and fast sites. Mutation of Ala-334 to a threonine in the slow site of the bovine type I regulatory subunit created a site with marked increase in cGMP affinity without changing cAMP affinity (Shabb, J. B., Ng. L., Corbin, J. D. (1990) J. Biol. Chem. 265, 16031-16034). The corresponding fast site residue (Ala-210) was changed to a threonine by oligonucleotide-directed mutagenesis, and a double mutant containing a threonine in each site was also made. Holoenzymes were formed from native catalytic subunit and each recombinant regulatory subunit. The fast site mutant holoenzyme exhibited an improved cGMP activation constant and an impaired cAMP activation constant. The double mutant cGMP/cAMP selectivity was 200-fold greater than that of wild-type holoenzyme, making it as responsive to cGMP as native cGMP-dependent protein kinase. The increased intrinsic binding energies of mutated sites for cGMP were 2.7-3.0 kcal mol-1, consistent with the presence of an extra hydrogen bond. Cyclic nucleotide analog studies implied that this hydrogen bond was between the threonine hydroxyl and the 2-amino of cGMP. Comparisons of amino acid sequences and cyclic nucleotide specificities suggested that the Ala/Thr difference may also impart cAMP/cGMP binding selectivity to related proteins such as cyclic nucleotide-gated ion channels.

Amino Acid Sequence↗