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50S is the larger subunit of the70Sribosome ofprokaryotes, i.e.bacteria andarchaea. It is the site of inhibition forantibiotics such asmacrolides,chloramphenicol,clindamycin, and thepleuromutilins. It includes the5S ribosomal RNA and23S ribosomal RNA.
Despite having the same sedimentation rate, bacterial and archaeal ribosomes can be quite different.
50S, roughly equivalent to the60S ribosomal subunit ineukaryotic cells, is the larger subunit of the70S ribosome of prokaryotes. The 50S subunit is primarily composed of proteins but also contains single-strandedRNA known asribosomal RNA (rRNA). rRNA forms secondary and tertiary structures to maintain the structure and carry out the catalytic functions of the ribosome.
X-ray crystallography has yieldedelectron density maps allowing the structure of the 50S inHaloarcula marismortui (archaeon) to be determined to 2.4Å resolution[1]and of the 50S in theDeinococcus radiodurans (bacterium) to 3.3Å.[2] The large ribosomal subunit (50S) is approximately twice as massive as the small ribosomal subunit (30S). The model of Hm 50S, determined in 2000 byNenad Ban and colleagues in the laboratory ofThomas Steitz and the laboratory ofPeter Moore, includes 2711 of the 2923nucleotides of 23SrRNA, all 122 nucleotides of its 5S rRNA, and structure of 27 of its 31proteins.[1]
Thesecondary structure of 23S is divided into six large domains, within which domain V is most important in itspeptidyl transferase[3] activity. Each domain contains normal secondary structure (e.g., base triple, tetraloop, cross-strand purine stack) and is also highly symmetric in tertiary structure; proteins intervene between their helices. Attertiary structure level, the large subunit rRNA is a single gigantic domain while the small subunit contains three structural domains. This difference reflects the lesser flexibility of the large subunit required by its function. While its core is conserved, it accommodates expansion segments on its periphery.[4][5]
AcryoEM structure of the 50S subunit from the archaeonMethanothermobacter thermautotrophicus has been determined. It shares the 50S size/sedimentation rate and the two rRNA count, but its 23S expansion segments have more in common with eukaryotes.[6]
A cryoEM reconstruction of the native 50S subunit of the extremely halophilic ArchaeanHalococcus morrhuae (classified underEuryarchaeota;Stenosarchaea group) is available. The 50S subunit contains a 108‐nucleotide insertion in its 5S rRNA,[7] which at subnanometer resolution, is observed to emerge from a four‐way junction without affecting the parental canonical 5S rRNA structure.[4]
Due to the differences, archaeal 50S are less sensitive to some antibiotics that target bacterial 50S.[8][9]
50S includes the activity that catalyzespeptide bond formation (peptidyl transfer reaction), prevents premature polypeptide hydrolysis, provides a binding site for the G-protein factors (assistsinitiation,elongation, and termination), and helpsprotein folding after synthesis.
An induced-fit mechanism has been revealed for how 50S catalyzes the peptidyl transfer reaction and prevents peptidyl hydrolysis. Theamino group of an aminoacyl-tRNA (binds to A site) attacks the carbon of acarbonyl group of a peptidyl-tRNA (binds to P site) and finally yields a peptide extended by oneamino acid esterified to the A site tRNA bound to the ribosomal A site and a deacylated tRNA in the P site.
When the A site is unoccupied, nucleotide U2620 (E. coli U2585), A2486 (2451) and C2106 (2063) sandwich the carbonyl group in the middle, forcing it into an orientation facing the A site. This orientation prevents anynucleophilic attack from the A site because the optimal attacking angle is 105 degrees from the plane of theester group. When a tRNA with a complete[?] CCA sequence at its acceptor stem is bound to the A site, C74 of the tRNA stacking with U2590 (2555) induces a conformational change in the ribosome, resulting in movement of U2541 (2506), U2620 (2585) through G2618 (2583). The displacement of bases allows the ester group to adopt a new conformation accessible to nucleophilic attack from the A site.
The N3 (nitrogen) of A2486 (2451) is closest to the peptide bond being synthesized and may function as a general base to facilitate the nucleophilic attack by the amino group of the aminoacyl-tRNA (in the A site). The pKa of A2486 (2451) is about 5 units higher in order tohydrogen bond with the amino group thus increasing its nucleophilicity. The elevation of pKa is achieved through a charge relay mechanism. A2486 (2451) interacts with G2482 (G2447), which hydrogen bonds with the buriedphosphate of A2486 (2450). This buried phosphate can stabilize the normally rare imino tautomers of both bases, resulting in an increase in the negative charge density on N3.
After initiation, elongation, and termination, there is a fourth step of the disassembly of the post-termination complex of ribosome,mRNA, and tRNA, which is a prerequisite for the next round of protein synthesis. The large ribosomal subunit has a role in protein folding bothin vitro andin vivo. The large ribosomal subunit provides ahydrophobic surface for the hydrophobic collapse step of protein folding. The newly synthesized protein needs full access to the large subunit to fold; this process may take a period of time (5 minutes forbeta-galactosidase[citation needed]).
The haloarchaea used in the current study were resistant to nalidixic acid, streptomycin, gentamicin, tetracycline, erythromycin, chloramphenicol, cephalothin, and clindamycin.