Yusaku Tagashira

An improved method for estimating sequence divergence between related DNAs from changes in restriction endonuclease cleavage sites.

SummaryWe have developed a theory to estimate the degree of sequence divergence between related DNAs from the comparison of restriction endonuclease recognition sites. Two major improvements have been made upon a similar method reported by Upholt (1977). First, the most probable value is calculated by the collective use of all available data. This reduces intrinsic statistical error and extends the analyzable range of sequence divergence. Second, all variables are redefined so that they have strict mathematical implications. This corrects a serious error arising from the misinterpretation of the meaning of the fraction of conserved cleavage sites. With this refined method, sequence divergence between rat and mouse mitochondrial DNAs (mtDNAs) was calculated to be about 25% substitutions/nucleotide, which is in good agreement with the DNA-DNA hybridization data obtained by Jakovcic et al. (1975). It was also estimated that the three types of rat mtDNAs differ from one another by 0.3 ~1% of total base pairs. These values are 2 ~5 times smaller than those obtained with the conventional method.

Evolutionary aspects of variant types of rat mitochondrial DNAs

Mitochondrial DNAs (mtDNAs) were prepared from various kinds of individual Norway rats, Rattus norvegicus, and from three types of individual black rats, Rattus rattus, (Asian type, Ceylon type, and Oceanian type). Intra- and interspecies divergence of their mtDNA sequences were calculated based on changes in restriction endonuclease cleavage sites. The extent of intraspecies divergence of black rats (about 8%) is much larger than that of Norway rats (1%) and the mtDNA of Asian-type black rats resembles the mtDNA of Norway rats more closely than it resembles the mtDNA of other types of black rats. These results strongly suggest that during the course of intraspecies differentiation of black rats, probably long after the separation of the three types of black rats, some Asian-type black rats were isolated sexually and formed a new species, Norway rats. On the basis of our observations we propose a hypothetical process to explain the evolution of animal mtDNA.

Two different molecular types of rat mitochondrial DNAs.

Abstract Experiments with restriction endonucleases showed that Sprague-Dawley strain rats have two types of mitochondrial DNAs (mtDNAs), named α- and β-types, which differ in primary structure. Individual rats have only one type. Individuals of other strains, such as Wistar, Fischer, and Donryu, also all have one of these types of mtDNA. Thus irrespective of their strain, all rats can be classified into one of two groups according to the type of their mtDNA.

Quantum-mechanical study on the photodimerization of aromatic molecules

Abstract Distinct parallelism is found between the quantum-mechanical index, the π delocalization energy and the degrees of dimer-forming ability of thymine, uracil and other aromatic compounds. It is surprising that the π delocalization energies for thymine and uracil are larger than those of well-known photodimer-forming compounds such as anthracene and acenaphthacene. The possible structures of thymine dimer isomers in vitro and in vivo , are postulated from considerations of, for example, the π delocalization energy and steric properties. Biological inactivation by photo-irradiation is discussed in relation to the energy liberation due to the thymine dimer formation.

The differences between the primary structures of mitochondrial DNAs from rat liver and ascites hepatoma (AH-130)

Rat mitochondrial DNAs (mtDNAs) of ascites hepatoma (AH-130) and normal liver cells (Donryu strain) were digested by various restriction endonucleases and the cleavage patterns compared by agarose gel electrophoresis. Different cleavage patterns were observed between AH-130 and liver mtDNAs when they were digested by HindII and EcoRI. The mtDNA of AH-130 lost one clevage site of HindII and one clevage site of EcoRI. The cleavage patterns of mtDNAs from other organs and strains tested were the same as that of liver mtDNA. From these observations we concluded that the molecular clone of AH-130 mtDNA was different from that of other mtDNAs.

Biochemical basis for the resistance of guinea pigs to carcinogenesis by 2-acetylaminofluorene

Abstract Studies were made on why guinea pigs are resistant to carcinogenesis by 2-acetylaminofluorene. Cytochrome P-448 and arylhydrocarbon hydroxylase were not induced in either the microsomes and nuclei of guinea pigs by 3-methylcholanthrene treatment. 3-Methylcholanthrene treatment caused only 2-fold increase in the binding of 2-acetylaminofluorene to DNA in nuclei isolated from guinea pigs, while it caused 17-fold increase in the binding in rat nuclei. Microsomes from 3-methylcholanthrene treated rats had 5 times more effect than Microsomes from 3-methylcholanthrene treated guinea pigs on the binding of 2-acetylaminofluorene to DNA of nuclei from untreated guinea pigs. N-Hydroxy-2-acetylaminofluorene combined equally well with the DNA of rats and guinea pigs. In guinea pigs, there was a good correlation between the low inducibility of cytochrome P-448 and the low binding of 2-acetylaminofluorene to DNA. Our results clearly showed that guinea pigs are resistant to tumor induction by 2-acetylaminofluorene through inability to carry out the first step of activation of 2-acetylaminofluorene.

A circular dichroism study on the conformation of DNA in rat liver chromatin.

Abstract 1. 1. The circular dichroism (CD) spectra of chromatins of rat liver and AH-130 hepatoma cells were measured, together with those in the presence of the reagents such as sodium dodecyl sulfate, NaCl and urea. Association of DNA with histone in chromatin was supposed to bring about deformation of DNA from the B form to the C form. 2. 2. On increasing the pH of the solution of chromatin, it was supposed that proteins were first disrupted and dissociated from chromatin. As a result, DNA would be subjected to the conformational change from the C form to the B form. The separation of the double helix was then expected. 3. 3. Conformation of DNA in chromatin of AH-130 hepatoma cells was substantially the same as that of normal rat liver.

The semi-empirical SCF Treatment of σ-π system I. Peptide molecule

Abstract The semi-empirical ASMO SCF method has been applied to the peptide molecule. In this treatment, σ and π electrons are explicitly taken into account. With regard to σ electrons, the molecular orbital originally used by Sandorfy (1955) and extensively developed by Fukui et al. (1962) in the Huckel MO method is employed, and the integrals involved in this treatment are estimated semi-empirically. The calculated absorption maximum, its oscillator strength, ionization potential and dipole moment are compared with experimental data and good agreement is found. The important role of the hydrogen atom in semiconductivity of protein and the possibility of σ-σ ∗ , π-σ ∗ and σ-π ∗ transitions of the peptide molecule are pointed out.

Sequence search on a supercomputer

A set of programs was developed for searching nucleic acid and protein sequence data bases for sequences similar to a given sequence. The programs, written in FORTRAN 77, were optimized for vector processing on a Hitachi S810-20 supercomputer. A search of a 500-residue protein sequence against the entire PIR data base Ver. 1.0 (1) (0.5 M residues) is carried out in a CPU time of 45 sec. About 4 min is required for an exhaustive search of a 1500-base nucleotide sequence against all mammalian sequences (1.2M bases) in Genbank Ver. 29.0. The CPU time is reduced to about a quarter with a faster version.

Length polymorphisms of restriction fragments of rat mitochondrial DNAs

Abstract Differences were found in the HpaII cleavage patterns of two types of rat ( Rattus norvegicus ) mtDNA, types A and B. One HpaII fragment, Hpa5, of type A was about 30 base pairs smaller than that of type B, but no 30-base pair fragment was detectable in an HpaII digest of type A mtDNA. Moreover, one HaeIII fragment, which is overlapped by Hpa5 in the cleavage map, was also about 30-base pairs smaller in type A than in type B. Thus, the length polymorphism of Hpa5 in the two types is probably not caused by HpaII site gain or loss, but by sequence deletion or insertion.