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

The carboxyl-terminal region of diphtheria toxin (DT) has been analysed in order to determine regions of receptor recognition. Biochemical cleavage of the toxin with hydroxylamine (HA) was used to generate the peptides HA9DT (residues 454-535), HA6DT (residues 482-535), and HA3DT (residues 454-481). Characterization of HA6DT demonstrated that the final 54 amino acids of DT are sufficient to constitute the receptor-binding domain of the toxin. Within HA9DT, the region encompassing HA3DT and containing the highly cationic polyphosphate-binding site did not contribute to the binding ability of HA6DT. Consistent with this observation, HA3DT itself did not compete for binding of radiolabelled DT to Vero cells. A 30-amino-acid synthetic peptide composed of residues 506-535 did not block receptor binding of DT, indicating that residues toward the amino-terminus of HA6DT, or the entire HA6DT region, are required for receptor recognition.
Mol Microbiol 1993 Feb
PMID:Characterization of the diphtheria toxin receptor-binding domain. 768 20

We have shown that deoxycytidine-5'-triphosphate modified by O-(4-aminobutyl)hydroxylamine in the pyrimidine ring, is effectively incorporated into DNA synthesizing in vitro, replacing deoxythymidine-5'-triphosphate or deoxycytidine-5'-triphosphate and inducing A-->G and G-->A transitions, respectively. UV spectroscopy and NMR spectroscopy have shown that the modified cytidine-5'-triphosphate is identical to N4-(4-aminobutoxy)-2'-deoxycytidine-5'-triphosphate. When the modified deoxycytidine-5'-triphosphate was inserted into DNA in vitro by DNA polymerase I of E. coli Klenow fragment, retardation sites correlating with poly-A sites (when the modified triphosphate replaced deoxythymidine-5'-triphosphate) or with poly-G sites (when it replaced deoxycytidine-5'-triphosphate) were revealed. Our data show high mutagenic effect of the modified deoxycytidine-5'-triphosphate inserted into DNA, allowing us to recommend this compound for localized static mutagenesis.
Mol Biol (Mosk)
PMID:[The mutagenic activity of N4-(4-aminobutoxy)-2'-deoxycytidine-5'-triphosphate]. 778 36

The cDNA encoding sea bass (Dicentrarchus labrax) prolactin (sbPRL) was obtained by reverse transcription-polymerase chain reaction (RT/PCR) from pituitary RNA with degenerate primers designed on the basis of the cDNAs of the two PRLs (tPRL188 and tPRL177) from the tilapia, Oreochromis niloticus. The sbPRL cDNA encodes a preprotein of 212 amino acids composed of a putative signal peptide of 24 residues and a mature protein of 188 amino acids that is the homologue of tiPRL188. The cDNA coding for the mature protein was cloned into the pAX4a+ expression vector and expressed efficiently in Escherichia coli as a beta-galactosidase-fusion protein. To split the fusion protein, a sequence encoding the hexapeptide, (Asn-Gly)3, that contains three Asn-Gly hydroxylamine-cleavable bonds, had been previously introduced by PCR upstream of the sbPRL cDNA. N-terminal sequencing confirmed that the cleaved product corresponded to sbPRL. An antiserum raised against the recombinant hormone detected by immunoblotting a single band in sea bass pituitaries and two bands in tilapia pituitaries, suggesting the occurrence of a single PRL form in sea bass.
Biochem Mol Biol Int 1994 Aug
PMID:The prolactin of European sea bass (Dicentrarchus labrax L.): cloning of cDNA and efficient expression in Escherichia coli. 780 37

We have carried out a genetic analysis of Escherichia coli HlyB using in vitro(hydroxylamine) mutagenesis and regionally directed mutagenesis. From random mutagenesis, three mutants, temperature sensitive (Ts) for secretion, were isolated and the DNA sequenced: Gly10Arg close to the N-terminus, Gly408Asp in a highly conserved small periplasmic loop region PIV, and Pro624Leu in another highly conserved region, within the ATP-binding region. Despite the Ts character of the Gly10 substitution, a derivative of HlyB, in which the first 25 amino acids were replaced by 21 amino acids of the lambda Cro protein, was still active in secretion of HlyA. This indicates that this region of HlyB is dispensable for function. Interestingly, the Gly408Asp substitution was toxic at high temperature and this is the first reported example of a conditional lethal mutation in HlyB. We have isolated 4 additional mutations in PIV by directed mutagenesis, giving a total of 5 out of 12 residues substituted in this region, with 4 mutations rendering HlyB defective in secretion. The Pro624 mutation, close to the Walker B-site for ATP binding in the cytoplasmic domain is identical to a mutation in HisP that leads to uncoupling of ATP hydrolysis from the transport of histidine. The expression of a fully functional haemolysin translocation system comprising HlyC,A,B and D increases the sensitivity of E. coli to vancomycin 2.5-fold, compared with cells expressing HlyB and HlyD alone. Thus, active translocation of HlyA renders the cells hyperpermeable to the drug. Mutations in hlyB affecting secretion could be assigned to two classes: those that restore the level of vancomycin resistance to that of E. coli not secreting HlyA and those that still confer hypersensitivity to the drug in the presence of HlyA. We propose that mutations that promote vancomycin resistance will include mutations affecting initial recognition of the secretion signal and therefore activation of a functional transport channel. Mutations that do not alter HlyA-dependent vancomycin sensitivity may, in contrast, affect later steps in the transport process.
Mol Gen Genet 1994 Nov 15
PMID:Identification and preliminary characterization of temperature-sensitive mutations affecting HlyB, the translocator required for the secretion of haemolysin (HlyA) from Escherichia coli. 780 92

Nitecapone [3-(3,4-dihydroxy-5-nitrobenzylidene)-2,4-pentanedione], is a scavenger of nitric oxide produced in vitro. It reduced the rate of methemoglobin formation from oxyhemoglobin exposed to nitric oxide generated from the reaction of hydroxylamine with Complex I of catalase and it decreased the amount of nitrite formed in the reaction of oxygen with nitric oxide generated from sodium nitroprusside. Nitecapone also affected the L-arginine dependent accumulation of nitrite in a suspension of peritoneal rat neutrophils. The related compounds entacapone [2-cyano-N, N-diethyl-3-(3,4 dihydroxy-5-nitrobenzyl)-propenamide] and OR 1246 [3-(3,4-dihydroxy-5-nitrobenzyl)-2,4-pentanedione] were also able to scavenge nitric oxide. The action of nitecapone on nitric oxide expands the role of nitecapone as a scavenger of reactive oxygen species, and suggests nitecapone, entacapone and OR 1246 as potential therapeutic agents for the treatment of diseases connected with increased production of nitric oxide.
Biochem Mol Biol Int 1994 Oct
PMID:Nitecapone: a nitric oxide radical scavenger. 783 30

The aim of this study was to investigate whether short-term treatment with nitric oxide donors could mimic cytokine inhibition of insulin secretion. We tested the nitric oxide generating compounds 3-morpholinosydnonimine (SIN-1), S-nitroso-N-penicillamine (SNAP), S-nitrosoglutathione and hydroxylamine for their ability to inhibit insulin secretion, raise cyclic GMP and lower cyclic AMP levels in isolated rat islets of Langerhans and the insulin-secreting cell lines HIT-T15 and RINm5F. In islets, all nitric oxide donors inhibited glucose-induced insulin secretion and raised cyclic GMP levels. SIN-1 and S-nitrosoglutathione also reduced cyclic AMP, while SNAP and hydroxylamine had no effect. Insulin secretion in HIT-T15 cells was inhibited by SIN-1, SNAP and hydroxylamine and in RINm5F cells by hydroxylamine. Inhibition of HIT-T15 and RINm5F cell insulin secretion was not accompanied by an increase in cyclic GMP levels. The degree of inhibition of insulin secretion was unrelated to the extent of release of nitric oxide by the compounds as measured by nitrite and nitrate production. More effective inhibition by S-nitrosoglutathione and hydroxylamine versus SIN-1 and SNAP may be related to intracellular versus extracellular site of nitric oxide generation.
Mol Cell Endocrinol 1994 Jun
PMID:The effect of nitric oxide donors on insulin secretion, cyclic GMP and cyclic AMP in rat islets of Langerhans and the insulin-secreting cell lines HIT-T15 and RINm5F. 792 70

Cardiac antiarrhythmic compounds are a diverse group divided into classes that differ in their mechanisms of action. Recent attention has focused on class III compounds, which prolong the action potential by blocking K+ channels. The purpose of this study was to characterize the mechanisms of actions of a class III compound, clofilium, and a simple analog, hydroxylamine, on an inactivating K+ channel. The defined system used a cloned inactivating K+ channel (Shaker-B) expressed in Xenopus oocytes. This channel is similar in physiological properties and core sequence to the inactivating K+ channel cloned from mammalian heart. Results presented here demonstrate that clofilium (100 microM) and hydroxylamine (10 mM) can cause use-dependent block, depending on the sequence of the pore region. A mutation of the pore known to influence selectivity and tetraethylammonium binding (threonine-441 to serine) confers use-dependent sensitivity to hydroxylamine and clofilium. Hybrid channels were formed from the coinjection of wild-type and mutant channel mRNAs; the analysis of block with the hybrid channels suggests that binding of hydroxylamine involves all subunits of the tetrameric channel, whereas clofilium affects channels containing as few as one mutant subunit. The simplest interpretation is that all four subunits contribute to an internal binding site for blockers such as clofilium and hydroxylamine and threonine-441 influences this binding site. The effectiveness of clofilium, unlike hydroxylamine, on the hybrid channels may reflect its structural complexity, which could allow interaction with a broader receptor site. Future studies will test this idea using other class III-related compounds.
Mol Pharmacol 1994 Nov
PMID:Block of the inactivating potassium channel by clofilium and hydroxylamine depends on the sequence of the pore region. 796 88

Hydroxylamine (HA) mutagenesis of an HA-induced splicing-defective bacteriophage T4 td intron mutant with a mutation in the intron P3 RNA pairing region was used to generate pseudorevertants. Because HA can only cause GC to AT transitions, the original mutant (H104A) could not undergo true reversion, yet the compensatory mutation on the opposite side of the P3 helix, which was complementary to the original H104A mutation, could occur. A pseudorevertant was isolated that contained both the original H104A mutation and the compensatory mutation HS9. By phenotypic and molecular genetic criteria, this double mutant (H104A-HS9) was shown to be able to undergo significant RNA splicing, thus confirming the existence and functional importance of the long-range P3 pairing region in this phage intron. The second-site suppressor mutation (HS9) was isolated by phage cross and also exhibited some self-splicing ability. A correlation exists between the strength of P3 helix Watson-Crick base pairing and the apparent level of splicing when wild-type, H104A, HS9, and H104A-HS9 are compared. This suggests that the primary role of the P3 RNA pairing region in the T4 td intron is structural in contributing to the critical RNA secondary structure.
Mol Microbiol 1994 Jul
PMID:A non-directed, hydroxylamine-generated suppressor mutation in the P3 pairing region of the bacteriophage T4 td intron partially restores self-splicing capability. 798 96

Arginine biosynthesis in Escherichia coli is negatively regulated by a hexameric repressor protein, encoded by the gene argR and the corepressor arginine. By hydroxylamine mutagenesis two types of argR mutants were isolated and mapped. The first type is transdominant. In heterodiploids, these mutant polypeptides reduce the activity of the wild-type repressor, presumably by forming heteropolymers. Four mutant repressor proteins were purified. Two of these map in the N-terminal half of the protein. Gel retardation experiments showed that they bind poorly to DNA, but they could be precipitated by L-arginine at the same concentration as the wild-type repressor. The other two mutant repressors map in the C-terminal half of the protein. They are poorly precipitated by L-arginine and they bind poorly to DNA. In addition, one of these mutants appears to exist as a dimer. The second type of argR mutant repressor consists of super-repressors. Such mutants behave as arginine auxotrophs as a result of hyper-repression of arginine biosynthetic enzymes. They map at many locations throughout the argR gene. Three arginine super-repressor proteins were purified. In comparison with the wild-type repressor, two of them were shown to have a higher DNA-binding affinity in the absence of bound arginine, while the third was shown to have a higher DNA-binding affinity when bound to arginine.
Mol Microbiol 1994 Aug
PMID:Mutational analysis of the arginine repressor of Escherichia coli. 799 72

The mechanism by which NAD stimulates cardiac adenylate cyclase was investigated. In highly purified canine cardiac sarcolemma, NAD stimulated adenylate cyclase activity in the presence of agents which activate Gs (i.e. 5 mM AlF4-, 10 microM GTP gamma S, 10 microM GppNHp or isoproterenol plus 2 nM GTP gamma S). Furthermore, the EC50 of isoproterenol to stimulate adenylate cyclase was reduced in the presence of NAD. In membranes incubated with [32P]-NAD, AlF4-, 10 microM GTP gamma S or isoproterenol plus 2 nM GTP gamma S produced a selective increase in the radiolabeling of a single 45-kDa protein which was identified as Gs alpha by immunoprecipitation. Cholera toxin catalysed radiolabeling of the same protein. Neutral hydroxylamine released [32P]-ADP-ribose from Gs alpha prelabeled in the presence of AlF4- and [32P]-NAD indicating that an arginine residue on Gs alpha was modified by an endogenous ADP-ribosyltransferase. ADP-ribosyltransferase inhibitors, novobiocin, vitamin K1 or 3-aminobenzamide, inhibited AlF4- stimulated ADP-ribosylation of Gs alpha and NAD potentiation of adenylate cyclase with similar efficacies. The activity responsible for NAD potentiation of adenylate cyclase and ADP-ribosylation of Gs alpha was not removed under hypotonic or hypertonic conditions and therefore appears to be tightly membrane bound. Collectively, these observations indicate that canine cardiac sarcolemma possess an ADP-ribosyltransferase which may constitutively catalyse transfer of an ADP-ribose to activated Gs alpha.
J Mol Cell Cardiol 1994 Feb
PMID:Modification of cardiac membrane adenylate cyclase activity and Gs alpha by NAD and endogenous ADP-ribosyltransferase. 800 86


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>