TRNA precursors

TFIIIC subunits TFIIIC90 and TFIIICllO. The known function of TFIIIC is to initiate transcription complex formation by binding to promoter DNA and recruiting TBP-containing TFIIIB and RNA polymerase III, that directs synthesis of tRNA precursors. TFIIIC have an intrinsic HAT activity. 

Ribonuclease II [EC 3.1.13.1], also called exoribo-nuclease II, catalyzes the exonucleolytic cleavage of the polynucleic acid, preferring single-stranded RNA, in the 3 - to 5 -direction to yield 5 -phosphomononucleotides. The enzyme processes 3 -terminal extra-nucleotides of monomeric tRNA precursors, following the action of ribonuclease P. Similar enzymes include RNase Q, RNase BN, RNase PHI, and RNase Y. Ribonuclease T2 [EC 3.1.27.1] is also known as ribonuclease II. 

This enzyme [EC 3.1.26.3], also known as RNase O and RNase D, catalyzes the endonucleolytic cleavage of RNA to 5 -phosphomonoesters. The enzyme cleaves multimeric tRNA precursors at the spacer region and is also involved in the processing of precursor rRNA, hnRNA, and early T7-mRNA. This enzyme can also act on double-stranded DNA. 

This system [EC 3.1.26.5] catalyzes the endonucleolytic cleavage of RNA, removing the 5 -extra-nucleotide from the tRNA precursor. This step is essential for tRNA processing. It generates the 5 -termini of mature tRNA... 

Transfer RNA precursors may undergo further posttranscriptional processing. The 3 -terminal trinucleotide CCA(3 ) to which an amino acid will be attached during protein synthesis (Chapter 27) is absent from some bacterial and all eukaryotic tRNA precursors and is added during processing (Fig. 26-23). This addition is carried out by tRNA nucleotidyltransferase, an unusual enzyme that binds the three ribonucleoside triphosphate precursors in separate active sites and catalyzes formation of the phosphodiester bonds to produce the CCA(3 ) sequence. The creation of this defined sequence of nucleotides is therefore not dependent on a DNA or RNA template—the template is the binding site of the enzyme. 

The best known processing nuclease is RNase P, which cleaves bacterial tRNA precursors to create the 5 ends of the mature tRNAs. All of the 64 tRNA precursors present in E. coli are cleaved by this unusual enzyme,221 222 which contains an essential piece of RNA (Chapter 12, Section D,6). Cleavage of polycis-tronic tRNA precursors by RNase P or of the previous-... 

Figure 28-10 Sequence of an E. coli tyrosine tRNA precursor drawn in a hypothetical secondary structure. Nucleotides found modified in the mature tRNA are indicated with their modifications (S4, 4-thiouridine Gm, 2 -0-methylgua-nosine 1°, N6-isopentenyladenosine jt, pseudouridine T, ribothymidine see also Fig. 5-33).241 A partial sequence of the tRNA gene past the CCA end is also shown. Note the region of local 2-fold rotational symmetry (indicated by the bars and the dot). The anticodon 3 -CUA (shaded) of this suppressor tRNA pairs with termination codon 5 -UAG.

Most eukaryotic tRNAs are formed from monomeric precursors, each gene acting as a transcriptional unit. Processing is similar to that in bacteria (Section A, 7). Eukaryotic RNase P usually cleaves the 5 end,555-558c and another enzyme cuts at the 3 end.542 556 The 3 CCA sequence of the mature tRNA is usually not present in the primary transcript but is added.559/559a b As in bacteria (p. 1620) extensive modification of bases also occurs in the tRNA precursors of eukaryotes.235 560-562 Many tRNA genes contain introns, which must be removed by splicing (Section 5). 

Figure 28-20 Composite structure representing several tRNA precursors arranged in a similar secondary structure (see also Fig. 5-30). The arrows indicate splice points. Variable positions are designated (O) for the mature tRNA and (X) for the intervening sequence and also in loops where insertions or deletions occur. From Ogden et al.603...

Figure 28-21 Two related splicing pathways for tRNA precursors. (A) Pathway followed in yeast. (B) Related pathway used by HeLa cells.598 602...

Pre-tRNAs. In the removal of type I introns the formation of specific stem and loop structures directs the splicing reactions (Fig. 28-18).47 597 Stems and loop structures already exist in tRNA precursors. Cleavage sites are usually located just to the 3 side of the anti-... 

The promoters for RNA polymerase III have been clearly proven to be internal on the template. In one instance the 5 S gene promoter lies between 45 and 83, whereas in tRNA genes it is split into two separate locations, one between 8 and 30 and a second between 51 and 72. Termination in 5 S genes is caused by a sequence of four A residues situated between two GC-rich regions. tRNA precursors are converted into mature tRNAs by a series of alterations (see later). 

Virtually all the initial products of transcription are further processed in eukaryotes. For example, tRNA precursors are... 

Figure 28.23. Transfer RNA Precursor Processing. The conversion of a yeast tRNA precursor into a mature tRNA requires the removal of a 14-nucleotide intron (yellow), the cleavage of a 5 leader (green), and the removal of UU and the attachment of CCA at the 3 end (red). In addition, several bases are modified.

RNAs also act as ribonucleases, cleaving other RNA molecules (e.g., RNase P cleaves tRNA precursors). 

RNase P cleaves a specific phosphodiester bond of tRNA precursors to form the mature 5 -end of tRNA. Bacterial RNase P is a natural occurring ribozyme composed of a catalytic RNA subunit and one or more proteins. Each bacterial RNAse P holoenzyme also consists of a small basic protein that stabilizes the folded structure of the RNA under physiological conditions. It may also help to discriminate pre-tRNA substrate from tRNA product and to mediate RNAse P dimerization. The three domains of life bacteria, Archaea, and Eukarya have similar structurally related RNAse P RNAs but different protein subunits this indicates that RNAse P is very ancient... 

The self-splicing introns and the RNA component of RNase P (which cleaves the 5 end of tRNA precursors) are two examples of ribozymes. These biological catalysts have the properties of true enzymes. They generally promote hydrolytic cleavage and transesterification, using RNA as substrate. Combinations of these reactions can be promoted by the excised group 1 intron of Tetrahymena rRNA, resulting in a type of RNA polymerization reaction. 

RNA (Chapfer 12, Section D,6). Cleavage of polycis-fronic tRNA precursors by RNase P or of the previous-... 

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