Molecular phylogeny of the ETS gene family. (1/263)

We have constructed a molecular phylogeny of the ETS gene family. By distance and parsimony analysis of the ETS conserved domains we show that the family containing so far 29 different genes in vertebrates can be divided into 13 groups of genes namely ETS, ER71, GABP, PEA3, ERG, ERF, ELK, DETS4, ELF, ESE, TEL, YAN, SPI. Since the three dimensional structure of the ETS domain has revealed a similarity with the winged-helix-turn-helix proteins, we used two of them (CAP and HSF) to root the tree. This allowed us to show that the family can be divided into five subfamilies: ETS, DETS4, ELF, TEL and SPI. The ETS subfamily comprises the ETS, ER71, GABP, PEA3, ERG, ERF and the ELK groups which appear more related to each other than to any other ETS family members. The fact that some members of these subfamilies were identified in early metazoans such as diploblasts and sponges suggests that the diversification of ETS family genes predates the diversification of metazoans. By the combined analysis of both the ETS and the PNT domains, which are conserved in some members of the family, we showed that the GABP group, and not the ERG group, is the one most closely related to the ETS group. We also observed that the speed of accumulation of mutations in the various genes of the family is highly variable. Noticeably, paralogous members of the ELK group exhibit strikingly different evolutionary speed suggesting that the evolutionary pressure they support is very different.  (+info)

An Lrp-like protein of the hyperthermophilic archaeon Sulfolobus solfataricus which binds to its own promoter. (2/263)

Regulation of gene expression in the domain Archaea, and specifically hyperthermophiles, has been poorly investigated so far. Biochemical experiments and genome sequencing have shown that, despite the prokaryotic cell and genome organization, basal transcriptional elements of members of the domain Archaea (i.e., TATA box-like sequences, RNA polymerase, and transcription factors TBP, TFIIB, and TFIIS) are of the eukaryotic type. However, open reading frames potentially coding for bacterium-type transcription regulation factors have been recognized in different archaeal strains. This finding raises the question of how bacterial and eukaryotic elements interact in regulating gene expression in Archaea. We have identified a gene coding for a bacterium-type transcription factor in the hyperthermophilic archaeon Sulfolobus solfataricus. The protein, named Lrs14, contains a potential helix-turn-helix motif and is related to the Lrp-AsnC family of regulators of gene expression in the class Bacteria. We show that Lrs14, expressed in Escherichia coli, is a highly thermostable DNA-binding protein. Bandshift and DNase I footprint analyses show that Lrs14 specifically binds to multiple sequences in its own promoter and that the region of binding overlaps the TATA box, suggesting that, like the E. coli Lrp, Lrs14 is autoregulated. We also show that the lrs14 transcript is accumulated in the late growth stages of S. solfataricus.  (+info)

The Bradyrhizobium japonicum nolA gene encodes three functionally distinct proteins. (3/263)

Examination of nolA revealed that NolA can be uniquely translated from three ATG start codons. Translation from the first ATG (ATG1) predicts a protein (NolA1) having an N-terminal, helix-turn-helix DNA-binding motif similar to the DNA-binding domains of the MerR-type regulatory proteins. Translation from ATG2 and ATG3 would give the N-terminally truncated proteins NolA2 and NolA3, respectively, lacking the DNA-binding domain. Consistent with this, immunoblot analyses of Bradyrhizobium japonicum extracts with a polyclonal antiserum to NolA revealed three distinct polypeptides whose molecular weights were consistent with translation of nolA from the three ATG initiation sites. Site-directed mutagenesis was used to produce derivatives of nolA in which ATG start sites were sequentially deleted. Immunoblots revealed a corresponding absence of the polypeptide whose ATG start site was removed. Translational fusions of the nolA mutants to a promoterless lacZ yielded functional fusion proteins in both Escherichia coli and B. japonicum. Expression of NolA is inducible upon addition of extracts from 5-day-old etiolated soybean seedlings but is not inducible by genistein, a known inducer of the B. japonicum nod genes. The expression of both NolA2 and NolA3 requires the presence of NolA1. NolA1 or NolA3 is required for the genotype-specific nodulation of soybean genotype PI 377578.  (+info)

The influence of C-terminal extension on the structure of the "J-domain" in E. coli DnaJ. (4/263)

Two different recombinant constructs of the N-terminal domain in Escherichia coli DnaJ were uniformly labeled with nitrogen-15 and carbon-13. One, DnaJ(1-78), contains the complete "J-domain," and the other, DnaJ(1-104), contains both the "J-domain" and a conserved "G/F" extension at the C-terminus. The three-dimensional structures of these proteins have been determined by heteronuclear NMR experiments. In both proteins the "J-domain" adopts a compact structure consisting of a helix-turn-helix-loop-helix-turn-helix motif. In contrast, the "G/F" region in DnaJ(1-104) does not fold into a well-defined structure. Nevertheless, the "G/F" region has been found to have an effect on the packing of the helices in the "J-domain" in DnaJ(1-104). Particularly, the interhelical angles between Helix IV and other helices are significantly different in the two structures. In addition, there are some local conformational changes in the loop region connecting the two central helices. These structural differences in the "J-domain" in the presence of the "G/F" region may be related to the observation that DnaJ (1-78) is incapable of stimulating the ATPase activity of the molecular chaperone protein DnaK despite evidence that sites mediating the binding of DnaJ to DnaK are located in the 1-78 segment.  (+info)

Crystal structure of the human Pax6 paired domain-DNA complex reveals specific roles for the linker region and carboxy-terminal subdomain in DNA binding. (5/263)

Pax6, a transcription factor containing the bipartite paired DNA-binding domain, has critical roles in development of the eye, nose, pancreas, and central nervous system. The 2.5 A structure of the human Pax6 paired domain with its optimal 26-bp site reveals extensive DNA contacts from the amino-terminal subdomain, the linker region, and the carboxy-terminal subdomain. The Pax6 structure not only confirms the docking arrangement of the amino-terminal subdomain as seen in cocrystals of the Drosophila Prd Pax protein, but also reveals some interesting differences in this region and helps explain the sequence specificity of paired domain-DNA recognition. In addition, this structure gives the first detailed information about how the paired linker region and carboxy-terminal subdomain contact DNA. The extended linker makes minor groove contacts over an 8-bp region, and the carboxy-terminal helix-turn-helix unit makes base contacts in the major groove. The structure and docking arrangement of the carboxy-terminal subdomain of Pax6 is remarkably similar to that of the amino-terminal subdomain, and there is an approximate twofold symmetry axis relating the polypeptide backbones of these two helix-turn-helix units. Our structure of the Pax6 paired domain-DNA complex provides a framework for understanding paired domain-DNA interactions, for analyzing mutations that map in the linker and carboxy-terminal regions of the paired domain, and for modeling protein-protein interactions of the Pax family proteins.  (+info)

Crystal structure of the Zalpha domain of the human editing enzyme ADAR1 bound to left-handed Z-DNA. (6/263)

The editing enzyme double-stranded RNA adenosine deaminase includes a DNA binding domain, Zalpha, which is specific for left-handed Z-DNA. The 2.1 angstrom crystal structure of Zalpha complexed to DNA reveals that the substrate is in the left-handed Z conformation. The contacts between Zalpha and Z-DNA are made primarily with the "zigzag" sugar-phosphate backbone, which provides a basis for the specificity for the Z conformation. A single base contact is observed to guanine in the syn conformation, characteristic of Z-DNA. Intriguingly, the helix-turn-helix motif, frequently used to recognize B-DNA, is used by Zalpha to contact Z-DNA.  (+info)

Putative mechanism of natural transformation as deduced from genome data. (7/263)

Genetic transformation is widely utilized in molecular biology as a tool for gene cloning in Escherichia coli and for gene mapping in Bacillus subtilis. Several strains of eubacteria can naturally take up exogenous DNA and integrate the DNA into their own genomes. Molecular details of natural transformation, however, remained to be elucidated. The complete genome of a cyanobacterium, Synechocystis sp. PCC6803, has been sequenced. This bacterium has been used to examine functions of a particular gene. The genome is considered to carry information on natural transformable characteristics of Synechocystis. The first step in genetic transformation is the uptake of exogenous DNA. Proteins with non-specific DNA binding features are required, because specificity in the exogenous DNA has not been demonstrated. Such proteins have modules interacting with the phosphate backbone of DNA, including helix-turn-helix modules. Using a consensus pattern of the phosphate-binding helix-turn-helix module, we searched through the genome data of Synechocystis for genes or open reading frame (ORF) products with the pattern in primary structures. We found that an ORF, slr0197, has the pattern in duplicate at the C-terminal region. We also found that the ORF product has a hydrophobic segment at the N-terminal region, which is followed by two internal repeats of the endonuclease domain. Based on these observations, we propose a model for the initial stage of genetic transformation. This is apparently the first report on molecular mechanisms of natural transformation.  (+info)

Change in conformation by DNA-peptide association: molecular dynamics of the Hin-recombinase-hixL complex. (8/263)

The Hin-DNA complex is a molecular complex formed by the C-terminal 52mer peptide of the Hin-recombinase and a synthetic 13-bp hixL DNA. The peptide has three alpha-helices, the second and third of which form the helix-turn-helix motif to bind to the major groove. Both termini of the peptide reside within the minor groove. Three molecular dynamics simulations were performed based on the crystal structure of the Hin-DNA complex: one for the free Hin peptide, one for the free hixL DNA, and one for the complex. Analyses of the trajectories revealed that the dynamic fluctuations of both the Hin peptide and the hixL DNA were lowered by the complex formation. The simulation supported the experimental observation that the N-terminus and the helix-turn-helix motif were critical for formation of the complex, but the C-terminus played only a supportive role in DNA recognition. The simulations strongly suggested that the binding reaction should proceed by the induced fit mechanism. The ion and solvent distributions around the molecules were also examined.  (+info)