Distribution of Pyrimidine Oligonucleotides in Strands L and H of Bacillus subtilis DNA

Abstract

The distribution of pyrimidine oligodeoxynucleotide clusters in L and H strands of Bacillus subtilis DNA separated by methylated albumin-Kieselguhr has been determined. Preparations of native and singlestranded DNA were degraded with diphenylamine in formic acid, and the released isostichs with the general formula of PyPn+1 were separated on DEAE-cellulose by chain length. Eleven isostichs were obtained for strands L and H in unequal proportions. Each isostich fraction was subfractionated by base composition on DEAE-cellulose at pH 3.0. 61 Pyrimidine oligonucleotide clusters were separated from the H strand and only 46 from the L strand. The findings show a higher degree of asymmetry between the strands in the distribution of cytosine-rich clusters as compared with thymine-rich clusters. The longest cytosine oligodeoxynucleotide present in both strands is of chain length 5. There is an unusually high distribution of thymine oligodeoxynucleotides of length 5-11. Up to chain length 6, the distribution of thymine oligodeoxynucleotides between the strands is about equal; from chain length 7 to 11 they occur predominantly in the H strand.Direct chemical analyses (1) and nucleotide composition of RNA transcripts synthesized in vitro show that complementary strands of DNA from bacteria exhibit various degrees of chemical asymmetry, namely, that the L strand is purine-rich and the H strand is pyrimidine-rich. For example, Bacillus subtilis DNA represents a type in which the chemical asymmetry between the strands involves both purines and pyrimidines, while in Escherichia coli, the asymmetry is limited to only guanine and cytosine. Studies on in vivo transcription patterns of these bacteria (3,4) show that the extent of asymmetric transcription can be correlated with the degree of bias in the distribution of purines and pyrimidines between the complementary strands. In B. subtilis, 85-90% of the messenger RNA (mRNA) and all the stable RNA components [ribosomal RNA (rRNA) and transfer RNA (tRNA)] are transcribed from the pyrimidine-rich H strand; in E. coli, the bias in transcription pattern is more limited, namely, 30-35% of mRNA and 30% of tRNA are transcribed from the purine-rich L strand (3, 4). It has been proposed that pyrimidine-rich clusters (particularly oligocytidylate sequences) may act as some sort of recognition sites involved in strand selection for asymmetric transcription by RNA polymerase (5). We were interested in ascertaining whether cytosine residues are indeed more frequently clustered in the H strands of microbial DNAs. Earlier investigations of the distribution of pyrimidine isostichs in native microbial DNA preparations demonstrated that oligocytidylate sequences of length 4 or 5 are very scarce, whereas oligothymidylate sequences are found in amounts often exceeding random predictions (6).In this paper, we present the distribution of pyrimidine oligonucleotide clusters in complementary chains of B. subtilis DNA using methylated albumin-Kieselguhr (MAK) columns for separation of the s...

Conformations and structural transitions in polydeoxynucleotides

“…In particular, the presence of localized homo(dA) • homo(dT) regions would be expected to perturb the B -A transition and perhaps even prevent the formation of the A conformation. This appears to be the case for several viral and bacterial AT-rich DNAs and also for the B. subtilis DNA which is known to contain long oligo(dA) and oligo(dT) segments (Rudner et al, 1972). In fact it was shown previously that these DNAs are undergoing a transition from the Bto a C-like form at lower r.h. and under identical salt conditions (Brahms et ah, 1973;Champed and Brahms, 1974).…”

“…Instead these DNAs usually undergo a transition to the C-like form as the humidity is decreased (Brahms et al, 1973). In addition, DNA with a moderate GC content, e.g., Bacillis subtilis DNA (44% GC) but with long oligo (dA), oligo (dT) sequences (Yamagashi and Takahashi, 1971;Rudner et al, 1972), does not adopt the A conformation at low humidity. This latter point suggests that the base sequence has an influence on DNA conformational transition.…”