a A u u u ggauuuaua1

usQuA u C uAGAuUuucuU

Figure 6.3. Secondary structure model for Caenorhabditis elegans mitochondrial 16S-like rRNA (see the legend to Fig. 6.1 for details and references).

The analysis of NMR structure of the internal loop in the E. coli 16S rRNA fragment formed with sequences 1404-1412/1488-1498 (see Fig. 6.1) has shown that it adopts a fully helical conformation with base pairs U1406:U1495 and C1407:G1494 and the three adenines (A1408, A1492, and A1493) stacked within the helix (Fourmi et al., 1996). On the other hand, it was noted that the bulges and the mismatches can considerably alter the conformation (e.g., groove dimensions) of neighboring helical regions and result in bending the structure.

There are three pseudoknot helices within the secondary structure of 16S rRNA shown in Figure 6.1: the helices 17-19/916-918, 505-507/524-526, and 570-571/865-866. They are highly conserved in the 16S-like rRNAs and may play an important role in organization of ribosome functional centers.

It is customary to divide the 16S rRNA secondary structure into four parts: three major domains, namely 5' domain, central domain and 3' major domain, and 3'-end minor domain. In many aspects these domains behave like autonomous structural units. The major domains of 16S rRNA are enclosed by longrange double helices: the helix 27-37/547-556 encloses, as a stem, the 5' domain, the helix 921-933/ 1384-1396 confines the 3' major domain, and the central domain is between these two helices. The sequence 912-920 and the pseudoknot helix 17-19/916-918 connect the three major domains. Interestingly, the two other 16S rRNA pseudoknot helices are positioned near the interdomain junctions.

These long-range base-pair interactions define also a core secondary structure that is universal among the 16S and 16S-like rRNAs (see Fig. 6.1). The universal core seems to comprise the most principal structural and functional part of the 16S-like rRNA molecules. Indeed, the vast majority of mutations altering ribosome activities are localized in the 16S rRNA universal core. Most of modified nucleotide residues are also clustered in the universal core (Brimacombe et al., 1993). The primordial ribosome is thought to consist of its rRNA core.

Comparative analysis of secondary structures of different 16S and 16S-like rRNAs reveals regions of variable size that interrupt the universal core (see Fig. 6.1). They are not evolutionary conserved and have been termed "variable" or "divergent" regions (as well as "expansion or contraction segments"). The positions in the E. coli 16S rRNA secondary structure where variable regions occur (Fig. 6.1) can be expanded or contracted in rRNAs of other organisms (compare with structures in Figs. 6.2 and 6.3). The role of variable regions in ribosome structure and function is unknown. Up to now no functionally meaning mutations have been found in the variable regions of 16S-like rRNAs. It is worth noting that the breaks in the polynucleotide chain of the discontinuous rRNA molecules (see above, Section 6.2) have been found to occur only in the variable regions.

6.3.2. Secondary Structure of the Large-Subunit rRNA

As one can see from Fig. 6.5, that demonstrates the example of eubacterial 23S rRNA, the general principles of organization of the small-subunit and large-subunit rRNA secondary structures are the same. The relative frequencies of Watson-Crick and non-Watson-Crick base-pairs in the 16S-like and 23S-like rRNAs are equal. Structure of several hairpin-loop fragments of bacterial 23S rRNA have been resolved with atomic resolution and it was shown that their single-stranded regions, just as in the case of the 16S rRNA, are well ordered.

The major difference between these two classes of molecules is that the 3' and 5' terminal sequences of 23S and 23S-like rRNAs, in contrast to the 16S-like rRNAs, are mutually complementary and form a long stable stem. In eukaryotic large-subunit rRNAs whose 5' terminal region is represented with 5.8S rRNA, the 3'-end sequence of 23S-like (25S or 28S) rRNA form a double-helical structure with the 5'-end sequence of the 5.8S rRNA, the latter being associated with the large rRNA due to formation of two more double helices. In chloroplasts the 5'-end sequence of the 23S rRNA forms a double-helical complex with the 3'-end region of the 4.5S rRNA.

The secondary structure of prokaryotic, eukaryotic, chloroplast and some mitochondrial 23S-like rRNAs consist of six domains (I-VI) enclosed by long range double helices. In the case of the E. coli 23S rRNA they are the helix 15-24/516-525 (domain I); the helix 579-584/1256-1261 (domain II); the helix 1295-1298/1642-1645 (domain III); the helix 1648-1667/1979-1988 (domain IV); the helix 2043-2057/ 2611-2625 (domain V), and the helix 2630-2644/2771-2788 (domain VI). There are approximately 15

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