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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The p160 coactivators, steroid receptor coactivator 1, glucocorticoid receptor interacting protein 1 (GRIP1) and the activator of thyroid and retinoic acid receptor, have two activation domains, AD1 and AD2, which transmit the activation signal from the DNA-bound nuclear receptor to the chromatin and/or transcription machinery. In screening for mammalian proteins that bind the AD2 of GRIP1, we identified a mouse actin-binding protein, alpha actinin 2 (mACTN2). mACTN2 was expressed in the heart, skeletal muscle, lung, brain and testis, but there was no expression in the spleen, liver or kidney. Interestingly, the expression level of mACTN2 in the developing embryo depended on the embryonic stage. We further demonstrated that mACTN2 could enhance two transactivation activities of GRIP1, which in turn could enhance the homodimerization of mACTN2. Importantly, mACTN2 not only served as a primary coactivator for androgen receptor, estrogen receptor and thyroid receptor activities, but also acted synergistically with GRIP1 to enhance these nuclear receptor (NR) functions. However, the NR binding motif, LXXLL, conserved in mACTN2 and other actinin family proteins, might be a dispensable domain for its coactivator roles in NRs. These findings suggested that mACTN2 might play an important role in GRIP1-induced NR coactivator functions.
J Mol Endocrinol 2004 Apr
PMID:The enhancement of nuclear receptor transcriptional activation by a mouse actin-binding protein, alpha actinin 2. 1507 53

To examine the substrate recognition mechanism of the human MTH1 protein, which hydrolyzes 2-hydroxy-dATP, 8-hydroxy-dATP, and 8-hydroxy-dGTP, ten nucleotide analogs (8-bromo-dATP, 8-bromo-dGTP, deoxyisoinosine triphosphate, 8-hydroxy-dITP, 2-aminopurine-deoxyriboside triphosphate, 2-amino-dATP, deoxyxanthosine triphosphate, deoxyoxanosine triphosphate, dITP, and dUTP) were incubated with the MTH1 protein. Of these, the former five nucleotides were hydrolyzed with various efficiencies. The fact that the syn-oriented brominated nucleotides were hydrolyzed suggests that the MTH1 protein binds to deoxynucleotides adopting the syn-conformation. However, 8-hydroxy-dITP, which lacks the 2-amino group of 8-hydroxy-dGTP, was degraded with tenfold less efficiency as compared with 8-hydroxy-dGTP. In addition, deoxyisoinosine triphosphate, lacking the 6-amino group of 2-hydroxy-dATP, was hydrolyzed as efficiently as 8-hydroxy-dGTP, but less efficiently than 2-hydroxy-dATP. These results clarify the effects of the anti/syn conformation and the functional groups on the 2 and 6 positions of the purine ring on the recognition by the human MTH1 protein.
J Mol Biol 2004 Feb 27
PMID:Probing the substrate recognition mechanism of the human MTH1 protein by nucleotide analogs. 1509 64

Mutants in deoxyadenosine methyltransferase (dam) from many Gram-negative pathogens suggest multiple roles for Dam methylase: directing post-replicative DNA mismatch repair to the correct strand, guiding the temporal control of DNA replication and regulating the expression of multiple genes (including virulence factors) by differential promoter methylation. Dam methylase (HI0209) in strain Rd KW20 was inactivated in Haemophilus influenzae strains Rd KW20, Strain 12 and INT-1; restriction with Dam methylation-sensitive enzymes DpnI and DpnII confirmed the absence of Dam methylation, which was restored by complementation with a single copy of dam ectopically expressed in cis. Despite the lack of increased mutation frequency, the dam mutants had a 2-aminopurine-susceptible phenotype that could be suppressed by secondary mutations in mutS, suggesting a role for Dam in H. influenzae DNA mismatch repair. Invasion of human brain microvascular endothelial cells (HBMECs) and human respiratory epithelial cells (NCI-H292) by the dam mutants was significantly attenuated in all strains, suggesting the absence of a Dam-regulated event necessary for uptake or invasion of host cells. Intracellular replication was inhibited only in the Strain 12 dam mutant, whereas in the infant rat model of infection, the INT-1 dam mutant was less virulent. Dam activity appears to be necessary for both in vitro and in vivo virulence in a strain-dependent fashion and may function as a regulator of gene expression including virulence factors.
Mol Microbiol 2004 Jul
PMID:Inactivation of deoxyadenosine methyltransferase (dam) attenuates Haemophilus influenzae virulence. 1522 41

By stopped-flow kinetics using 2-aminopurine as a probe to detect base flipping, we show here that base flipping by the Escherichia coli Dam DNA-(adenine-N6)-methyltransferase (MTase) is a biphasic process: target base flipping is very fast (k(flip)>240 s(-1)), but binding of the flipped base into the active site pocket of the enzyme is slow (k=0.1-2 s(-1)). Whereas base flipping occurs in the absence of S-adenosyl-l-methionine (AdoMet), binding of the target base in the active site pocket requires AdoMet. Our data suggest that the tyrosine residue in the DPPY motif conserved in the active site of DNA-(adenine-N6)-MTases stacks to the flipped target base. Substitution of the aspartic acid residue of the DPPY motif by alanine abolished base flipping, suggesting that this residue contacts and stabilizes the flipped base. The exchange of Ser188 located in a loop next to the active center by alanine led to a seven- to eightfold reduction of k(flip), which was also reduced with substrates having altered GATC recognition sites and in the absence of AdoMet. These findings provide evidence that the enzyme actively initiates base flipping by stabilizing the transition state of the process. Reduced rates of base flipping in substrates containing the target base in a non-canonical sequence demonstrate that DNA recognition by the MTase starts before base flipping. DNA recognition, cofactor binding and base flipping are correlated and efficient base flipping takes place only if the enzyme has bound to a cognate target site and AdoMet is available.
J Mol Biol 2004 Aug 06
PMID:Stopped-flow and mutational analysis of base flipping by the Escherichia coli Dam DNA-(adenine-N6)-methyltransferase. 1527 35

Heterogeneous ribonucleoprotein A1 (hnRNP A1) is a prototype for the family of eukaryotic RNA processing proteins containing the common RNA recognition motif (RRM). The region consisting of residues 1-195 of hnRNP A1 is referred to as UP1. This region has two RRMs and has a high affinity for both single-stranded RNA and the human telomeric repeat sequence d(TTAGGG)(n). We have used UP1's novel DNA binding to investigate how RRMs bind nucleic acid bases through their highly conserved RNP consensus sequences. Nine complexes of UP1 bound to modified telomeric repeats were investigated using equilibrium fluorescence binding and X-ray crystallography. In two of the complexes, alteration of a guanine to either 2-aminopurine or nebularine resulted in an increase in K(d) from 88nM to 209nM and 316nM, respectively. The loss of these orienting interactions between UP1 and the substituted base allows it to flip between syn and anti conformations. Substitution of the same base with 7-deaza-guanine preserves the O6/N1 contacts but still increases the K(d) to 296nM and suggests that it is not simply the loss of affinity that gives rise to the base mobility, but also the stereochemistry of the specific contact to O6. Although these studies provide details of UP1 interactions to nucleic acids, three general observations about RRMs are also evident: (1) as suggested by informatic studies, main-chain to base hydrogen bonding makes up an important aspect of ligand recognition (2) steric clashes generated by modification of a hydrogen bond donor-acceptor pair to a donor-donor pair are poorly tolerated and (3) a conserved lysine position proximal to RNP-2 (K(106)-IFVGGI) orients the purine to allow stereochemical discrimination between adenine and guanine based on the 6-position. This single interaction is well-conserved in known RRM structures and appears to be a broad indicator for purine preference in the larger family of RRM proteins.
J Mol Biol 2004 Sep 17
PMID:Human UP1 as a model for understanding purine recognition in the family of proteins containing the RNA recognition motif (RRM). 1534 34

Despite sharing many common features, adenine-binding and guanine-binding sites in proteins often show a clear preference for the cognate over the non-cognate ligand. We have analyzed electrostatic potential (ESP) patterns at adenine and guanine-binding sites of a large number of non-redundant proteins where each binding site was first annotated as adenine/guanine-specific or non-specific from a survey of primary literature. We show that more than 90% of ESP variance at the binding sites is accounted for by only two principal component ESP vectors, each aligned to molecular dipoles of adenine and guanine. Projected on these principal component vectors, the adenine/guanine-specific and non-specific binding sites, including adenine-containing dinucleotides, show non-overlapping distributions. Adenine or guanine specificities of the binding sites also show high correlation with the corresponding electrostatic replacement (cognate by non-cognate ligand) energies. High correlation coefficients (0.94 for 35 adenine-binding sites and 1.0 for 20 guanine-binding sites) were obtained when adenine/guanine specificities were predicted using the replacement energies. Our results demonstrate that ligand-free protein ESP is an excellent indicator for discrimination between adenine and guanine-specific binding sites and that ESP of ligand-free protein can be used as a tool to annotate known and putative purine-binding sites in proteins as adenine or guanine-specific.
J Mol Biol 2004 Sep 17
PMID:Electrostatic potential of nucleotide-free protein is sufficient for discrimination between adenine and guanine-specific binding sites. 1534 56

An orphan G protein-coupled receptor from rat has recently been discovered to be activated by the nucleobase adenine (Proc Natl Acad Sci USA 99:8573-8578, 2002). In the present study, we show for the first time that the adenine receptor is expressed in membrane preparations of native tissues and cell lines in high density, including rat brain cortex, rat brain striatum, and the mouse neuroblastoma x rat glioma hybrid cell line NG108-15. Saturation analysis with [3H]adenine at rat brain cortical membranes exhibited a single high-affinity binding site with a KD value of 27.2 nM, and a binding capacity of 2.28 pmol/mg of protein. Kinetic studies revealed unusual binding kinetics of [3H]adenine with rapid association and slow dissociation. A series of compounds were investigated in [3H]adenine competition experiments at rat brain cortex. Only minor substitution of the adenine structure was tolerated, the most potent compounds of the present series being 2-fluoroadenine (Ki value of 620 nM), 8-thioadenine (Ki value of 2.77 microM), N6-methyladenine (Ki value of 3.64 microM), and 7-methyladenine (Ki value of 4.13 microM), all of which were partial agonists (40-60% intrinsic activity). Adenine dose dependently inhibited forskolin-stimulated adenylate cyclase in membrane preparations of NG108-15 cells as well as in intact cells, showing that the receptor is functional in NG108-15 cells. Reverse transcriptase-polymerase chain reaction experiments followed by sequencing indicate that the NG108-15 cells express the murine ortholog of the adenine receptor. Moreover, preliminary radioligand binding studies with [3H]adenine at membranes of human astrocytoma 1321N1 cells suggest that a human ortholog of the rat adenine receptor exists.
Mol Pharmacol 2005 Mar
PMID:Evidence for the functional expression and pharmacological characterization of adenine receptors in native cells and tissues. 1560 13

The Zap1 transcription factor is a central player in zinc homeostasis in yeast. This protein regulates the expression of genes involved in zinc accumulation and storage. For most of its target genes, Zap1 activates expression in zinc-limited cells and this function is inhibited in replete cells. Zap1 has two activation domains, AD1 and AD2, which are independently regulated by zinc status. In this study, we characterized AD1 and its regulation by zinc. AD1 was mapped using deletions to residues 332-402 of Zap1. The region required for the zinc responsiveness of this activation domain, designated 'ZRD(AD1), was mapped to residues 182-502. Thus, AD1 is embedded within its larger zinc-responsive domain. Using a combination of in silico analysis, random mutagenesis and site-directed mutagenesis, we identified key residues within ZRD(AD1) required for its regulation by zinc. Most of these residues are cysteines and histidines that could potentially serve as Zn(II) ligands. These results suggest that ZRD(AD1) senses zinc by direct Zn(II) binding. Consistent with this hypothesis, purified ZRD(AD1) bound multiple Zn(II) ions. Finally, our results indicate that, in the context of the full-length Zap1 protein, AD1 and AD2 are both critical to the full control of gene expression in response to zinc.
Mol Microbiol 2005 Aug
PMID:Zap1 activation domain 1 and its role in controlling gene expression in response to cellular zinc status. 1604 25

Like in bacteria, DNA in these organisms is subjected to enzymatic modification (methylation) both at adenine and cytosine residues. There is an indirect evidence that adenine DNA methylation takes place also in animals. In plants m6A was detected in total, mitochondrial and nuclear DNAs; in plants one and the same gene (DRM2) can be methylated both at adenine and cytosine residues. ORF homologous to bacterial adenine DNA-methyltransferases are present in nuclear DNA of protozoa, yeasts, insects, nematodes, higher plants, vertebrates and other eukaryotes. Thus, adenine DNA-methyltransferases can be found in the various evolutionary distant eukaryotes. First N6-adenine DNA-methyltransferase (wadmtase) of higher eukaryotes was isolated from vacuolar fraction of vesicles obtained from aging wheat coleoptiles; in the presence of S-adenosyl-L-methionine this Mg2+ -, Ca2+ -dependent enzyme de novo methylates first adenine residue in TGATCA sequence in single- and double-stranded DNA but it prefers single-stranded DNA structures. Adenine DNA methylation in eukaryotes seems to be involved in regulation of both gene expression and DNA replication including replication of mitochondrial DNA. It can control persistence of foreign DNA in a cell and seems to be an element of R-M system in plants. Thus, in eukaryotic cell there are, at least, two different systems of the enzymatic DNA methylations (adenine and cytosine ones) and a special type of regulation of gene functioning based on the combinatory hierarchy of these interdependent genome modifications.
Mol Biol (Mosk)
PMID:[Methylation of adenine residues in DNA of eukaryotes]. 1608 5

Ty1 retrotransposons of the yeast Saccharomyces cerevisiae are activated by different kinds of stress. Here we show that Ty1 transcription is stimulated under severe adenine starvation conditions. The Bas1 transcriptional activator, responsible for the induction of genes of the de novo AMP biosynthesis pathway (ADE) in the absence of adenine, is not involved in this response. Activation occurs mainly on Ty1 elements, whose expression is normally repressed by chromatin and is suppressed in a hta1-htb1Delta mutant that alters chromatin structure. Activation is also abolished in a snf2Delta mutant. Several regions of the Ty1 promoter are necessary to achieve full activation, suggesting that full integrity of the promoter sequences might be important for activation. Together, these observations are consistent with a model in which the activation mechanism involves chromatin remodeling at Ty1 promoters. The consequence of Ty1 transcriptional activation in response to adenine starvation is an increase in Ty1 cDNA levels and a relief of Ty1 dormancy. The retrotransposition of four native Ty1 elements increases in proportion to their increase in transcription. Implications for the regulation of Ty1 mobility by changes in Ty1 mRNA levels are discussed.
Mol Cell Biol 2005 Sep
PMID:Severe adenine starvation activates Ty1 transcription and retrotransposition in Saccharomyces cerevisiae. 1610 95


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