Eukaryotic DNA does not form nucleosomes as readily as some prokaryotic DNA.

Nucleosomal-length DNA was prepared from the genomic DNA of various prokaryotic and eukaryotic organisms by limited nuclease digestion after reconstitution with core histones. The DNAs ranged in base composition from 26.5% to 72% guanosine-plus-cytosine (%GC). The nucleosomal-length DNAs were then used in a competitive reconstitution assay in order to quantitatively determine their relative abilities to form nucleosomes. The results of the assay indicate a linear dependence of the free energy of nucleosome formation on base composition and, surprisingly, show that several prokaryotic DNAs form nucleosomes as well as or better than eukaryotic DNAs.     

Compositional Heterogeneity and Patterns of Molecular Evolution in the Drosophila Genome

The rates and patterns of molecular evolution in many eukaryotic organisms have been shown to be influenced by the compartmentalization of their genomes into fractions of distinct base composition and mutational properties. We have examined the Drosophila genome to explore relationships between the nucleotide content of large chromosomal segments and the base composition and rate of evolution of genes within those segments. Direct determination of the G + C contents of yeast artificial chromosome clones containing inserts of Drosophila melanogaster DNA ranging from 140-340 kb revealed significant heterogeneity in base composition. The G + C content of the large segments studied ranged from 36.9% G + C for a clone containing the hunchback locus in polytene region 85, to 50.9% G + C for a clone that includes the rosy region in polytene region 87. Unlike other organisms, however, there was no significant correlation between the base composition of large chromosomal regions and the base composition at fourfold degenerate nucleotide sites of genes encompassed within those regions. Despite the situation seen in mammals, there was also no significant association between base composition and rate of nucleotide substitution. These results suggest that nucleotide sequence evolution in Drosophila differs from that of many vertebrates and does not reflect distinct mutational biases, as a function of base composition, in different genomic regions. Significant negative correlations between codon-usage bias and rates of synonymous site divergence, however, provide strong support for an argument that selection among alternative codons may be a major contributor to variability in evolutionary rates within Drosophila genomes.     

Determination of DNA base composition by small scale acrylamide-CsCl gradient centrifugation

In this paper we describe a simple and rapid protocol for DNA base composition determination by CsCl gradient in the presence of acrylamide. This method permits the determination of GC content in microgram amounts of DNA, and results are easily documented in photographs or graphs. The protocol was applied to the characterization of nematode DNA, but can be used for other organisms. Analyzing several experiments the mean standard deviation observed in the calculated GC content is near 1.3%.     

Characteristics of glutamate dehydrogenase, a new diagnostic marker for the genus Fusobacterium

Enzymes representative of carbohydrate and nitrogen metabolism were screened for their presence and activity amongst species of the genus Fusobacterium. Glutamate dehydrogenase (GDH) was reliably detected in all 25 strains studied. The pH profile of this enzyme and the DNA base composition of selected strains were also determined. DNA base composition of selected strains ranged between 28-32.9 mol% G + C. GDH was active between pH 7.5-11.5 but two pH profiles of activity, with optima at 9.5 and 10.5, were discernible among species. Apart from Fusobacterium nucleatum, which had a heterogeneous enzyme pattern, the GDH electrophoretic mobility was constant within a species but in a few cases the enzyme bands overlapped. A combination of the pH profile, the GDH electrophoretic pattern and the DNA base composition provided clear separation of the test organisms into discrete groups; however, a larger number of strains must be examined before the full potential of these tests can be evaluated.     

Co-polymer tracts in eukaryotic, prokaryotic, and organellar DNA.

Large variations in DNA base composition and noticeable strand asymmetries are known to occur between different organisms and within different regions of the genomes of single organisms. Apparently such composition and sequence biases occur to fulfill structural rather than informational requirements. Here we report the wide occurrence of a more subtle biasing of DNA sequence that can have structural consequences: an increase or a suppression of the number of long tracts of two-base co-polymers. Strong biases were observed when the DNA sequences of the longest eukaryotic, prokaryotic, and organellar entries in the GenBank data base (totaling 773 kilobases) were analyzed for the number of occurrences of tracts of the two-base co-polymers (A,T)n, (G,C)n, and (A,C)n as a function of tract length. (The expression (A,T)n is used here to denote an uninterrupted tract, n nucleotides in length, of A and T bases in any proportion or order, terminated at each end by a G or C residue.) Characteristic differences are also observed in tract biases of eukaryotic vs. prokaryotic organisms.     

Relative roles of primary sequence and (G + C)% in determining the hierarchy of frequencies of complementary trinucleotide pairs in DNAs of different species

To an approximation Chargaff's rule (%A = %T; %G = %C) applies to single-stranded DNA. In long sequences, not only complementary bases but also complementary oligonucleotides are present in approximately equal frequencies. This applies to all species studied. However, species usually differ in base composition. With the goal of understanding the evolutionary forces involved, I have compared the frequencies of trinucleotides in long sequences and their shuffled counterparts. Among the 32 complementary trinucleotide pairs there is a hierarchy of frequencies which is influenced both by base composition (not affected by shuffling the order of the bases) and by base order (affected by shuffling). The influence of base order is greatest in DNA of 50% G + C and seems to reflects a more fundamental hierarchy of dinucleotide frequencies. Thus if TpA is at low frequency, all eight TpA-containing trinucleotides are at low frequency. Mammals and their viruses share similar hierarchies, with intra- and intergenomic differences being mainly associated with differences in base composition (percentage G + C). E. coli and, to a lesser extent, Drosophila melanogaster hierarchies differ from mammalian hierarchies; this is associated with differences both in base composition and in base order. It is proposed that Chargaff's rule applies to single-stranded DNA because there has been an evolutionary selection pressure favoring mutations that generate complementary oligonucleotides in close proximity, thus creating a potential to form stem-loops. These are dispersed throughout genomes and are rate-limiting in recombination. Differences in (G + C)% between species would impair interspecies recombination by interfering with stem-loop interactions.     

Hyperthermophiles and the problem of DNA instability

Rates of chemical decomposition of DNA at the optimal growth temperatures of hyperthermophiles seem incongruent with the requirements of accurate genome replication. The peculiar physiology, ecology and phylogeny of hyperthermophiles combine to suggest that these prokaryotes have solved a molecular problem (spontaneous loss of native DNA structure) of a magnitude that well-studied microorganisms do not face. The failure of DNA base composition to correlate with optimal growth temperature among hyperthermophiles provides indirect evidence that other mechanisms maintain their chromosomal DNA in the duplex form. Studies in vitro indicate that DNA primary structure is more difficult to maintain at extremely high temperature than is secondary structure, yet hyperthermophiles exhibit only modest levels of spontaneous mutation. Radiation sensitivity studies also indicate that hyperthermophiles repair their DNA efficiently in vivo , and underlying mechanisms are beginning to be examined. Several enzymes of DNA metabolism from hyperthermophilic archaea exhibit unusual biochemical features that may ultimately prove relevant to DNA repair. However, genomic sequencing results suggest that many DNA repair genes of hyperthermophilic archaea may not be recognized because they are not sufficiently related to those of well-studied organisms.     

Ultraviolet irradiation and repair--possible factors determining the evolution of DNA structure

It is suggested that DNA in first organisms on the Earth was enriched by GC-pairs due to intensive solar UV-irradiation. In the course of progressive evolution, nucleotide composition of DNA shifted to the favour of AT-pairs. This shift to higher photosensitivity became possible because of the development of reparation systems and screening of DNA in cells. The latter phenomenon is better revealed in higher organisms, accounting their stable TA-type of DNA. In lower forms, peculiarities of their habitats provided some conditions for variability of nucleotide composition. High content of GC-pairs remained in those of them which are subjected to the influence of solar irradiation. Parasitic microorganisms and those safely protected from solar radiation developed towards the AT-type of DNA. Both in higher and lower organisms, AT-pairs are richer in those zoms of DNA which are presented by several copies and in which mutations are less significant for the organism therefore being permissible. Structural genes exhibit heterogeneous nucleotide composition which provides for higher variability of the synthesized proteins.     

Dispersity of repeat DNA sequences in Oncopeltus fasciatus,an organism with diffuse centromeres

DNA of Oncopeltus fasciatus, an organism with diffuse centromeres, has been characterized by determination of its base composition, buoyant density, thermal stability, and reassociation kinetics; renatured DNA was characterized similarly. We conclude that repeated sequences are primarily short and scattered throughout the genome. This is in contrast to the extensive tandem repeats which are found in DNAs of organisms with discrete centromeres.     

Deoxyribonucleic acid base compositions of hyphomicrobia

Summary The deoxyribonucleic acids of 70 hyphomicrobia were examined at equilibrium in neutral CsCl density gradients. The guanine plus cytosine (%G+C) content was estimated to range from 59.2 to 66.8% G+C. The strains could be divided into three groups with different base composition of their DNA; 61.0±1.1%, 64.1±0.6%, and 66.5±0.6% G+C. The values are compared with those for the base composition of DNA of a number of phototrophic, budding bacteria.     

Directional mutation pressure,mutator mutations,and dynamics of molecular evolution

Using a general form of the directional mutation theory, this paper analyzes the effect of mutations in mutator genes on the G + C content of DNA, the frequency of substitution mutations, and evolutionary changes (cumulative mutations) under various degrees of selective constraints. Directional mutation theory predicts that when the mutational bias between A/T and G/C nucleotide pairs is equilibrated with the base composition of a neutral set of DNA nucleotides, the mutation frequency per gene will be much lower than the frequency immediately after the mutator mutation takes place. This prediction explains the wide variation of the DNA G + C content among unicellular organisms and possibly also the wide intragenomic heterogeneity of third codon positions for the genes of multicellular eukaryotes. The present analyses lead to several predictions that are not consistent with a number of the frequently held assumptions in the field of molecular evolution, including belief in a constant rate of evolution, symmetric branching of phylogenetic trees, the generality of higher mutation frequency for neutral sets of nucleotides, the notion that mutator mutations are generally deleterious because of their high mutation rates, and teleological explanations of DNA base composition. Presented at the NATO Advanced Research Workshop onGenome Organization and Evolution, Spetsai, Greece, 16–22 September 1992     

Classification of coryneform bacteria associated with human urinary tract infection (group D2) as Corynebacterium urealyticum sp. nov.

Urealytic strains of coryneform bacteria that are designated Corynebacterium group D2 and are isolated from human urine are a cause of urinary tract infections. Cell wall and lipid analyses confirmed that these organisms are members of the genus Corynebacterium but can be separated from other species in the genus on the basis of DNA base composition and DNA-DNA hybridization values. Biochemically, strains in this taxon can be distinguished from other Corynebacterium spp. by their failure to produce acid from carbohydrates, by their failure to reduce nitrates, and by their ability to hydrolyze urea. We regard these bacteria as a new species of the genus Corynebacterium and propose the name Corynebacterium urealyticum. The type strain is strain NCTC 12011 (= ATCC 43042).     

Characterization and Relatedness of Marine Vibrios Pathogenic to Fish: Deoxyribonucleic Acid Homology and Base Composition

Deoxyribonucleic acid (DNA) isolated from several pathogenic marine vibrios was utilized in studies of base composition and nucleotide sequence relationships. Patterns of relatedness inferred from DNA criteria were found to correlate closely with those inferred from morphological, physiological, and serological analysis of the same organisms. The utility of the phenotypic and genotypic approaches to the determination of relatedness is discussed.     

Quantitative Aspects of Deoxyribonucleic Acid Renaturation: Base Composition, State of Chromosome Replication, and Polynucleotide Homologies

The base composition of a deoxyribonucleic acid (DNA) sample affects its intrinsic rate of renaturation. In agreement with the information of Wetmur and Davidson, it was established that high guanosine plus cytosine (GC) DNA renatures faster than expected from analytical measurement of its molecular weight. A calculated correction factor of 1.8% of the observed C(0)t(.5) is required for every mole per cent GC difference from 51% GC. The correction factor is now established in the range of 32 to 65% GC. Renaturation of DNA mixtures prepared from pairs of organisms has been studied. When no similarity existed between the two organisms, the observed C(0)t(.5) of the mixture was the sum of the independently determined C(0)t(.5) values. Lack of additivity was correlated with similarities in polynucleotide sequence of the reassociating DNA molecules. A quantitative relationship was formulated to relate C(0)t(.5) values of renatured DNA mixtures to per cent binding ("homology"). Finally, it was demonstrated that DNA prepared from log-phase cells renatures faster than stationary-phase DNA and also departs from theoretical second-order kinetics.     

Deoxyribonucleic Acid Base Sequence Homologies of Some Budding and Prosthecate Bacteria

The genetic relatedness of a number of budding and prosthecate bacteria was determined by deoxyribonucleic acid (DNA) homology experiments of the direct binding type. Strains of Hyphomicrobium sp. isolated from aquatic habitats were found to have relatedness values ranging from 9 to 70% with strain "EA-617," a subculture of the Hyphomicrobium isolated by Mevius from river water. Strains obtained from soil enrichments had lower values with EA-617, ranging from 3 to 5%. Very little or no homology was detected between the amino acid-utilizing strain Hyphomicrobium neptunium and other Hyphomicrobium strains, although significant homology was observed with the two Hyphomonas strains examined. No homology could be detected between prosthecate bacteria of the genera Rhodomicrobium, Prosthecomicrobium, Ancalomicrobium, or Caulobacter, and Hyphomicrobium strain EA-617 or H. neptunium LE-670. The grouping of Hyphomicrobium strains by their relatedness values agrees well with a grouping according to the base composition of their DNA species. It is concluded that bacteria possessing cellular extensions represent a widely diverse group of organisms.     

Deoxyribonucleic acid base composition of species ofKlebsiella,Azotobacter andBacillus

Methods of molecular taxonomy were used to study the genome or deoxyribonucleic acid (DNA) of different strains of five genera of nitrogen-fixing bacteria. The DNA base compositions of all strains ofKlebsiella pneumoniae (previously classified as strains ofAchromobacter sp.,Aerobacter aerogenes orK. pneumoniae) occupy a fairly narrow range of values from 56.7 to 62.5% G-C content. No significant difference was observed in the DNA base composition of bacteria which can fix molecular nitrogen and that of strains which do not fix nitrogen.Six strains of the speciesAzotobacter vinelandii and the one strain ofA. chroococcum tested possess similar DNA base composition. The strain ofAzotobacter agilis tested, however, had a DNA base composition different from these seven strains. The G-C content of the strains ofAzotomonas insolita falls within the broad range of thePseudomonas genus (55–70%), but the peritrichous flagellation of these strains eliminates them from this genus. Their classification still remains to be ascertained.Eleven strains of the speciesBacillus polymyxa andB. macerans formed two homogenous groups of organisms with different DNA base composition. The atypical strainB. polymyxa Hino is genetically similar to the strains of the speciesBacillus macerans and perhaps should be reclassified as such.Supported in part by National Science Foundation grant GB-483 and National Institutes of Health grants AI 01417 (11) and based on Ph.D. thesis of senior author (1967).     

Entropy of the genetic information and evolution

The entropy of the amino acid sequences coded by DNA is considered as a measure of diversity or variety of proteins, and is taken as a measure of evolution. The DNA or m-RNA sequence is corsidered as a stationary second-order Markov chain composed of four kinds of bases. Because of the biased nature of the genetic code table, increase of entropy of amino acid sequences is possible with biased nucleotide sequence. Thus the biased DNA base composition and the extreme rarity of the base doubletC _p G of higher organisms are explained. It is expected that the amino acid composition was highly biased at the days of the origin of the genetic code table, and the more frequent amino acids have tended to get rarer, and the rarer ones more frequent. This tendency is observed in the evolution of hemoglobin, cytochrome C, fibrinopeptide, immunoglobulin and lysozyme, and protein as a whole.     

Cluster analysis of genes in codon space

Summary We construct a codon space in which a given DNA sequence can be plotted as a function of its base composition in each of the three codon positions. We demonstrate that the base composition is very highly nonrandom, with sequences from more primitive organisms having the least random compositions. By using cluster analysis on the points plotted in codon space we show that there is a strong correlation between base composition and type of organism, with the most primitive organisms having the highest A or T content in the second and third codon positions. A smooth transition toward lower A+T and higher G+C content is observed in the second and third codon positions as the evolutionary complexity of the organism increases. Besides this general trend, more detailed structure can be observed in the clustering that will become clearer as the data base is increased.     

DNA base composition of some sheathed bacteria

The DNA base composition of five recently isolated Haliscomenobacter hydrossis strains were compared with those of Sphaerotilus natans, Leptothrix cholodnii and Leptothrix discophora. The DNA base composition of H. hydrossis strains ranged from 48.3 to 49.7% GC, whereas the % GC values of S. natans, L. cholodnii and L. discophora were found to be 69.7, 69.6 and 71.2, respectively. These results indicate that Haliscomenobacter species and the microorganisms of the Sphaerotilus-Leptothrix group should not be classified into the same genus, as they are genetically at best remotely related.The reallocation of some Leptothrix species into the genus Sphaerotilus is not in contradiction with their DNA base composition.     

INTERSPECIFIC TRANSFORMATION IN BACILLUS

Marmur, J. (Brandeis University, Waltham, Mass.), E. Seaman, and J. Levine. Interspecific transformation in Bacillus. J. Bacteriol. 85:461-467. 1963.-Deoxyribonucleic acids (DNA) from various species of the taxonomic group Bacillaceae were examined for base composition, ability to carry out interspecific transformation, and formation of molecular hybrids in vitro. The minimal requirement for genetic compatibility among different species and for DNA interaction (both reflecting base sequence homologies) is the similarity of the guanine plus cytosine contents of the DNA. The close correlation between the ability of DNA to be competent in interspecific transformation and to form hybrid molecules on denaturation and annealing provided a rational approach to the study of genetic relationship among organisms for which no genetic exchange has yet been demonstrated. Any or all of the criteria (base composition of DNA, transformation, molecular hybrid formation) can be used as tools in the taxonomic assessment of closely related microorganisms.