A program for the identification of tRNA-like structures in DNA sequence data.

A computer algorithm has been developed which identifies tRNA genes and tRNA-like structures in DNA sequences. The program searches the sequence string for specific base positions that correspond to the invariant and semi-invariant bases found in tRNAs. The tRNA nature of the sequence is confirmed by the presence of complementary base pairing at the tRNA's calculated 5' and 3' ends (which in situ constitutes the amino-acyl stem region). The program achieves greater than 96% accuracy when run against known tRNA sequences in the Genbank database. The program is modular and is readily modified to allow searching either a file or database. The program is written in "C" and operates on a D.E.C. Vax 750. The utility of the algorithm is demonstrated by the identification of a distinctive tRNA structure in an intron of a published bovine hemoglobin gene.     

Dependence of M1 RNA substrate specificity on magnesium ion concentration.

We have constructed a plasmid expressing E. coli M1 RNA, the catalytic RNA subunit of ribonuclease P, under the control of a phage T7 promoter. The active M1 RNA species synthesized in vitro by T7 RNA polymerase from this vector was reacted with the tRNA(Gln) - tRNA(Leu) precursor RNA (Band K) encoded by phage T4. Only the tRNA(Leu) moiety of this dimeric precursor RNA contains the 3' terminal C-C-A sequence common to all tRNAs. We observed that protein-free M1 RNA was capable of processing the precursor RNA at the 5' ends of both tRNA tRNA sequences. The rate of cleavage of the tRNA(Gln) sequence was more strongly dependent on [Mg2+] than that of tRNA(Leu), increasing severalfold between 100 and 500 mM Mg2+, conditions under which the rate of cleavage at the tRNA(Leu) sequence was constant.     

Escherichia coli 16S rRNA 3''-end formation requires a distal transfer RNA sequence at a proper distance.

The 16S rRNA species in bacterial precursor rRNAs is followed by two evolutionarily conserved features: (i) a double-stranded stem formed by complementary sequences adjacent to the 5' and 3' ends of the 16S rRNA; and (ii) a 3'-transfer RNA sequence. To assess the possible role of these features, plasmid constructs with precursor-specific features deleted were tested for their capacity to form mature rRNA. Stem-forming sequences were dispensable for both 5' and 3' terminus formation; whereas an intact spacer tRNA positioned greater than 24 nucleotides downstream of the 16S RNA sequence was required for correct 3'-end maturation. These results suggest that spacer tRNA at an appropriate location helps form a conformation obligate for pre-rRNA processing, perhaps by binding to a nascent binding site in preribosomes. Thus, spacer tRNAs may be an obligate participant in ribosome formation.