Showing papers by "Harry F. Noller published in 1978"

Complete nucleotide sequence of a 16S ribosomal RNA gene from Escherichia coli.

Abstract: The complete nucleotide sequence of the 16S RNA gene from the rrnB cistron of Escherichia coli has been determined by using three rapid DNA sequencing methods. Nearly all of the structure has been confirmed by two to six independent sequence determinations on both DNA strands. The length of the 16S rRNA chain inferred from the DNA sequence is 1541 nucleotides, in close agreement with previous estimates. We note discrepancies between this sequence and the most recent version of it reported from direct RNA sequencing [Ehresmann, C., Stiegler, P., Carbon, P. & Ebel, J.P. (1977) FEBS Lett. 84, 337-341]. A few of these may be explained by heterogeneity among 16S rRNA sequences from different cistrons. No nucleotide sequences were found in the 16S rRNA gene that cannot be reconciled with RNase digestion products of mature 16S rRNA.

Altered topography of 16S RNA in the inactive form of Escherichia coli 30S ribosomal subunits.

Abstract: We have studied the topography of 16S RNA in the inactive form of the 30S ribosomal subunit (Ginsburg, I., et al. (1973) J. Mol. Biol. 79, 481), using the guanine-specific reagent kethoxal. Oligonucleotides surrounding reactive guanine residues were isolated and quantitated by means of diagonal electrophoresis and sequenced. Comparison of these results with experiments on active or reactivated subunits reveals the following: (1) Most of the sites which are reactive in active 30S subunits are much more reactive (average 13-fold) in inactive subunits. Upon reactivation, these sites return to a less reactive state. Thus, a reversible increase in accessibility of specific 16S RNA sites parallels the reversible loss of protein synthesis activity of 30S subunits. (2) The number of kethoxal-reactive sites in inactive subunits is about twice that of active subunits. The nucleotide sequences and locations of the additional accessible sites in inactive subunits have been determined. (3) Sites that can be located in the 16S RNA sequence are distributed throughout the RNA chain in inactive subunits, in contrast to the clustering observed in active subunits. (4) The sites of kethoxal substitution are single stranded. Yet, of the 30 sites that can be located, 23 were predicted to be base paired in the proposed secondary structure model for 16S RNA (Ehresmann, C., et al. (1975), Nucleic Acids Res. 2, 265).

Nucleotide sequences of accessible regions of 23S RNA in 50S ribosomal subunits.

Abstract: Nucleotide sequences around kethoxal-reactive guanine residues of 23S RNA in 50S ribosomal subunits have been determined. By use of the diagonal paper electrophoresis method )Noller, H.F. (1974), Biochemistry 13, 4694-4703), 41 ribonuclease T1 oligonucleotides, originating from about 25 sites, were identified and sequenced. These sites are single stranded and accessible in free 50S subunits, and are thus potential sites for interaction with functional ligands during protein synthesis. Examination of these sequences for potential intermolecular base-pairing reveals the following: (1) There are 19 possible complementary combinations between exposed sequences in 16S and 23S RNA containing more than 4 base pairs: 15 containing 5 base pairs and 4 containing 6 base pairs. Nine of these complementary combinations contain 16S RNA sequences which we have previously shown to be protected from kethoxall by 50S subunits (Chapman, N.M., and Noller, H.F. (1977), J. Mol. Biol. 109, 131-149). (2) One of the exposed sites in 23S RNA has a sequence which is complementary to the invariant GT psi CR sequence in tRNA.