A statistical survey of the torsion angles, bond angles, and bond lengths in the sugar and phosphate groups of well-refined mononucleoside, mononucleotide, dinucleoside monophosphate, and trinucleoside diphosphate crystal structures contained in the Cambridge Structural Database and the Nucleic Acid Database is reported. The mean values of the geometrical parameters in these structures and their estimated standard deviations are separated according to their chemistry and conformation. These new parameters serve as a basis for a dictionary of standard nucleic acid geometry.

Structures at atomic resolution (up to 1.0,&) which contain bases, sugars or the phosphodiester linkage, were selected from the Nucleic Acid Database or the Cambridge Structural Database to build a nucleic acid dictionary from X-ray refined structures. The dictionary consists of the average values for bond distances, bond angles and dihedral angles. The variance of the sample is used to provide information about the expected r.m.s, deviations of the refined parameters. A dictionary was constructed for refinement trials in X-PLOR. The dictionary includes RNA and DNA in C2'-endo and C3'-endo sugar pucker conformations, as well as values for the backbone dihedrals. Tests were performed on the dictionary using three structures: a B-DNA, a Z-DNA and a protein-DNA complex. During the course of refinement, all three structures showed significant improvements as measured by r.m.s, deviations and R factors when compared to the previous DNA dictionary.

We present estimates of the bond-length and bond-angle parameters for the nitrogenous base side groups of nucleic acids. These values are the result of a statistical survey of small molecules in the Cambridge Structural Database for which high-resolution X-ray and neutron crystal structures are available. The statistics include arithmetic means and standard deviations for the different samples, as well as comparisons of the population distributions for sugar-and non-sugar-derivatized bases. These accumulated data provide appropriate target values for refinements of oligonucleotide structures, as well as sets of standard atomic coordinates for the five common bases.

The classification feast͞famine regulatory proteins (FFRPs) encompasses archaeal DNA-binding proteins with Escherichia coli transcription factors, the leucine-responsive regulatory protein and the asparagine synthase C gene product. In this paper, we describe two forms of the archaeal FFRP FL11 (pot0434017), both assembled from dimers. When crystallized, a helical cylinder is formed with six dimers per turn. In contrast, in solution, disks are formed, most likely consisting of four dimers each; an observation by cryoelectron microscopy. Whereas each dimer binds a 13-bp sequence, different forms will discriminate between promoters, based on the numbers of repeating 13-bp sequences, and types of linkers inserted between them, which are either of 7-8 or Ϸ18 bp. The amino acid sequences of these FFRPs are designed to form the same type of 3D structures, and the transition between their assembly forms is regulated by interaction with small molecules. These considerations lead us to propose a possible mechanism for regulating a number of genes by varying assembly forms and by combining different FFRPs into these assemblies, responding to environmental changes.

Feast/famine regulatory proteins comprise a diverse family of transcription factors, which have been referred to in various individual identifications, including Escherichia coli leucine-responsive regulatory protein and asparagine synthase C gene product. A full length feast/famine regulatory protein consists of the N-terminal DNA-binding domain and the C-domain, which is involved in dimerization and further assembly, thereby producing, for example, a disc or a chromatin-like cylinder. Various ligands of the size of amino acids bind at the interface between feast/famine regulatory protein dimers, thereby altering their assembly forms. Also, the combination of feast/famine regulatory protein subunits forming the same assembly is altered. In this way, a small number of feast/famine regulatory proteins are able to regulate a large number of genes in response to various environmental changes. Because feast/famine regulatory proteins are shared by archaea and eubacteria, the genome-wide regulation by feast/famine regulatory proteins is traceable back to their common ancestor, being the prototype of highly differentiated transcription regulatory mechanisms found in organisms nowadays.

The crystal structure of TATA binding protein (TBP) from a mesothermophilic archaeon, Sulfolobus acidocaldarius, has been determined at a resolution of 2.0 A with an R factor of 20.9%. By comparing this structure with the structures of TBPs from a hyperthermophilic archaeon and mesophilic eukaryotes, as well as by comparing amino acid sequences of TBPs from archaea, covering a wide range of optimum growth temperatures, two significant determinants of the stability of TBP have been identified: increasing the interior hydrophobicity by interaction between three residues, Val, Leu, and Ile, with further differentiation of the surface, and increasing its hydrophilicity and raising the cost of unfolding. These findings suggest directions along which the stability of TBP can be engineered.

Histidine residues added to the N-terminus of a polypeptide (i.e. a His-tag) was used, for the first time to our knowledge, for electron labeling of the protein upon its electron spectroscopic imaging. Originally such a His-tag was developed by another group to purify modified proteins by taking advantage of their affinity to nickel. The feast/famine regulatory protein pot0434017 (FL11) was modified by adding six His residues to its N-terminus, so that each His pair would chelate a nickel ion. An electron microscope was operated at 200 KeV, and the electrons that lost the energy by ~875 eV upon interaction with the metal were selectively focused. The majority, 60–70%, of the spots detected in the electron micrographs were paired by distances shorter than 80 Å, and over 70% of them were paired by distances shorter than 40 Å. It is concluded that the protein molecules formed dimers, and the termini of most of the protein molecules were labeled with nickel by this method.

Abstract:Particles formed by a feast/famine regulatory protein (FFRP), pot0434017 (FL11), in solution in the absence of DNA were analyzed using electron microscopy (EM). By applying conventional (i.e. dry) EM to the protein negatively stained with uranyl acetate, top views of tetrameric assemblies of dimers were obtained, where four pairs each of N-domains were extending from C-domains assembled around the centers. In cryo-EM images of the protein embedded in 3D amorphous ice, sets of four densities were arranged around ellipsoids having similar lengths for their long axes but of different lengths for their short axes. These images were interpreted as projections with different tilts of four pairs of N-domains arranged inside flat assemblies: the positively charged N-domains only were stained with ammonium molybdate, but the negatively charged C-domains were unstained and thus unobservable. Using seventeen such cryo-images, in combination with a crystal structure equivalent to an assembly of C-domains, a disk-like 3D structure was reconstructed.