Ribosome

A ribosome is a small, dense granular particle of RNA and protein occurring both in prokaryotic as well as in eukaryotic cells as the protein synthesizing machinery that translates messenger RNA (mRNA) into a polypeptide chain of amino acids delivered by transfer RNA (tRNA). It can be thought of as a giant enzyme that builds a protein from a set of genetic instructions. Ribosomes occur either freely in the matrix of mitochondria, chloroplast, and cytoplasm (the internal fluid of the cell) or in membrane bound state in the endoplasmic reticulum and the nuclear envelope. Since ribosomes are ribozymes, it is thought that they might be remnants of the RNA world.

Ribosomes were first clearly described by Romanian cell biologist George Palade in the mid–1950s as dense particles or granules of ribonucleoprotein after observing under the electron microscope (Palade 1955) for which he would win the Nobel Prize. The term ribosome was later proposed by the scientist Richard B. Roberts in 1958 while writing the introductory comments for the symposium proceeding "Microsomal Particles and Protein Synthesis" (Roberts 1958).

The structure and function of the ribosomes and associated molecules, known as the translational apparatus, has been of research interest since the mid 20^th century and the focus of the study has been to work out the topology (shape and positions of the individual protein and rRNA) of ribosomes.

File:Ribosome subunits.png

Figure 1: Ribosome structure indicating small subunit (A) and large subunit (B). Side and front view.
(1) Head. (2) Platform. (3) Base. (4) Ridge. (5) Central protuberance. (6) Back. (7) Stalk. (8) Front.

Occurrence

The ribosomes occur both in prokaryotic as well as eukaryotic cells and both in plant as well as animal cell. An Escherichia coli cell contains 10,000 ribosomes, forming about 25 percent of the total bacterial cell mass. A cultured mammalian cell may contain as much as 10 million ribosomes. In prokaryotic cells, the ribosomes are distributed freely in the cytoplasm. In eukaryotic cells, they are found either freely floating in the matrix of mitochondria, chloroplast and cytoplasm or attached to the membrane of the endoplasmic reticulum and the nuclear envelope. Free and membrane–bound ribosomes differ only in their spatial distribution; they are identical in structure and function. Whether the ribosome exists in a free or membrane–bound state depends on the presence of an ER targeting signal sequence on the protein being synthesized.

Free ribosomes

Free ribosomes are "free" to move about anywhere in the cytoplasm (within the cell membrane). The yeast cells, reticulocytes or lymphocytes, meristematic plant tissues, embryonic nerve cells, and cancerous cells contain large number of free ribosomes. Proteins made by free ribosomes are used within the cell. Thus, the cells which synthesize specific proteins for the intracellular utilization and storage often contain large number of free ribosomes. Such cells are the erythroblasts, developing muscle cells, skin cells, and so forth.

Membrane–bound ribosomes

When certain proteins are synthesized, they need be "membrane–bound". Therefore, the new polypeptide chains are usually synthesized in membrane bound ribosomes and are inserted directly into the endoplasmic reticulum, from where they are then transported to their destinations. Bound ribosomes usually produce proteins that are used within the cell membrane or are expelled from the cell via exocytosis. Thus, in the cells actively engaged in protein synthesis, the ribosomes remain attached to the membranes of the endoplasmic reticulum. Such cells are the pancreatic cells, plasma cells, hepatic parenchymal cells, Nissls bodies, osteoblasts, serous cells or submaxillary gland cells, chief cells of the glandular stomach, thyroid cells, and mammary gland cells.

Structure

Overview

The various ribosomes share a core structure that is quite similar despite the large differences in size. Ribosomes are oblate spheroid granules with a diameter ranging from 15 to 25 nm (150 to 250 Å). Each ribosome is porous, hydrated, and consists of two subunits (Figure 1). One ribosomal subunit is larger in size and has a dome–like shape, while the other ribosomal subunit is smaller and occurs above the larger one forming a cap–like structure (Figure 2). The ribosomes are chemically composed of mainly RNA (ribosomal RNA, rRNA) and proteins (ribonucleoprotein, RNP). Lipid is totally absent in ribosomes. Both constituents occur approximately in equal proportion in its two subunits.

Crystallographic work has shown that there are no ribosomal proteins close to the reaction site for polypeptide synthesis. This suggests that the protein components of ribosomes act as a scaffold that may enhance the ability of rRNA to synthesize protein rather than directly participating in catalysis. Also it has been pointed out that all of the catalytic activity of the ribosome is carried out by the RNA, the proteins reside on the surface and seem to stabilize the structure (Alberts et al. 2002). (See: Ribozyme)

The two ribosomal subunits remain fitted together due to high concentration of Mg⁺⁺ ions. In the decreased Mg⁺⁺ concentration, the two subunits dissociate. Actually, in bacterial cells, the two subunits are found to occur freely in the cytoplasm and they come together only for the process of protein synthesis. At high concentration of Mg⁺⁺ ions in the matrix, two ribosomes (each called monosomes) become associated with each other and form what is known as dimer. Further, during the process of protein synthesis, many ribosomes are woven together in a common mRNA just like beads; the resulting structure is known as polyribosome or polysome.

Figure 2 : Large (1) and small (2) subunit fit together

Prokaryotic Ribosomes

Prokaryotes have comparatively smaller ribosomes with the sedimentation coefficient of 70 Svedberg unit (abbreviated as S), and a molecular weight of 2.7x10⁶ daltons. Each of the 70S ribosomes consists of a small (30S) and a large (50S) subunits. The 70S ribosomes contain proportionally more RNA than protein. For example, the ribosomes of E. coli contain 63 percent rRNA and 37 percent protein. The 70S ribosomes have three different types of rRNA, viz., 23S rRNA, 16S rRNA, and 5S rRNA. The large subunit is composed of a 5S rRNA subunit (consisting of 120 nucleotides), a 23S rRNA subunit (with 2900 nucleotides), and 34 proteins. The 30S subunit has a 16S rRNA subunit (with 1540 nucleotides) bound to 21 proteins (Alberts et al. 2002).

Eukaryotic Ribosomes

Eukaryotes have bigger ribosomes of 80S sedimentation coefficient and of 40x10⁶ daltons molecular weight. Each 80S ribosome consists of a small (40S) and large (60S) subunits. The ribosomal subunits of prokaryotes and eukaryotes are quite similar (Alberts et al. 2002). However, 80S ribosomes are composed of proportionally less RNA and more protein. For example, in pea seedling, ribosomes have 40 percent rRNA and 60 percent protein. There are four different types of rRNA in 80S ribosomes, viz., 28S rRNA (but 25–26S rRNA in plants, fungi, and protozoans), 18S rRNA, 5S rRNA, and 5.8S rRNA. The large 60S subunit is composed of a 5S RNA (120 nucleotides), a 28S RNA (4700 nucleotides), a 5.8S RNA (160 nucleotides) subunits and ~49 proteins. The 40S subunit has a 18S RNA (1900 nucleotides) subunit and ~33 proteins (Alberts et al. 2002). About 60 percent of the rRNA is helical (i.e., double stranded) and contains paired bases. These double stranded regions are due to hairpin loops between complimentary regions of the linear molecule. Thus we can say that the extra RNA in the larger ribosomes is in several long continuous insertions, such that they form loops out of the core structure without disrupting or changing it (Alberts et al. 2002).

The ribosomes found in chloroplasts and mitochondria of eukaryotes also consist of large and small subunits bound together into one 55S particle (Alberts et al. 2002). These organelles are believed to be descendants of bacteria (see Endosymbiotic theory) and as such their ribosomes are similar to those of prokaryotes (Alberts et al. 2002). The 55S ribosomes of mammalian mitochondria lack 5S rRNA, but contain 21S and 12S rRNAs. The 21S rRNA occurs in larger or 35S ribosomal subunit, while 12S rRNA occurs in smaller or 25S ribosomal subunit.

The differences between the prokaryotic and eukaryotic ribosomes are exploited by pharmaceutical chemists to create antibiotics that can destroy a bacterial infection without harming the cells of the infected person. Due to the differences in their structures, the bacterial 70S ribosomes are vulnerable to these antibiotics (such as Chloramphenicol) while the eukaryotic 80S ribosomes are not. Even though mitochondria possess ribosomes similar to the bacterial ones, mitochondria are not affected by these antibiotics because they are surrounded by a double membrane that does not easily admit these antibiotics into the organelle (O'Brien 1971).

Ultra–structure

Figure 3 : Molecular structure of the 50S Subunit. Proteins are shown in blue and RNA in orange.

The general molecular structure of the ribosome has been known since the early 1970s. In the early 2000s the structure has been achieved at high resolutions, in the order of a few angstroms and the papers giving the structure of the ribosome at atomic resolution were published in rapid succession in late 2000.

The structure of the 30S small subunit from Thermus thermophilus (a highly thermophilic bacteria first discovered in deep–sea hot vents) shows that the decoding center, which positions mRNA and three tRNAs, is constructed entirely of RNA (Schluenzen et al. 2000; Wimberly 2000). The mRNA threads through a tunnel within the small subunit. The 3’ end of the 16S rRNA is supposed to involve in mRNA binding. Each of the three tRNAs is bound in a distinctive binding sites made from structural elements contributed by both the 50S subunit and the 30S subunit. In each of the three tRNA binding sites (A–, P–, and E–sites), the ribosome contacts all of the major elements of tRNA, providing an explanation for the conservation of tRNA structure (Yusupov et al. 2001). The anticodon stem–loops of tRNAs point into the 30S subunit, whereas 3’ ends attached to amino acid or peptide through an acyl bond point down in 50S subunit. Further, the 3’ ends of the A–site and P–site tRNAs are juxtapositioned in the peptidyl transferase site of the 50S subunit. On the contrary, a metal ion stabilizes a kink in the mRNA that demarcates the boundary between A and P sites, which is potentially important to prevent slippage of mRNA, while translocation of tRNA from site A to P. Also metal ions stabilize the inter–subunit interface (Selmer 2006).

The 16S rRNA of the small, 30S subunit folds into four domains: 5', central, 3' major, and 3' minor. The structural autonomy of these domains implies that they move relative to one another. Thus, head of the small subunit shows high degree of flexibility compared to its rest of the body. Swiveling of the head observed by Schuwirth et al. (2005) suggests a mechanism for the final movements of messenger RNA (mRNA) and transfer RNAs (tRNAs) during translocation. Structural changes correlating events at the particle's far end with the cycle of mRNA translocation at the decoding region are transmitted by extended RNA helical elements that run longitudinally through its body (Schluenzen et al. 2000).

The 23S rRNA of the large, 50S subunit folds into six secondary structural domains, while the seventh domain is formed by 5S rRNA. The 50S subunit from the archea, Haloarcula marismortui is shown to have all its rRNAs to fit together like the pieces of a three–dimensional jigsaw puzzle to form a large, monolithic structure. Proteins are found everywhere on its surface except in the active site where peptide bond formation takes place and where it contacts the small subunit. Most of the proteins stabilize the structure by interacting with several rRNA domains (Ban et al. 2000). The large ribosomal subunit catalyzes peptide bond formation and binds initiation, termination, and elongation factors. The peptidyl transferase function is attributed to the 23S rRNA, making this RNA a ‘ribozyme’. Nascent polypeptides emerge through a tunnel in the large ribosome subunit. The tunnel lumen is lined with RNA helices and some ribosomal protein.

Biogenesis

Ribosomes are not self–replicating particles. Synthesis of various components of ribosomes such as rRNAs and proteins are under genetic control. In bacteria, a single gene transcript containing the sequences of 16S, 23S, and 5S rRNAs is synthesized by a rRNA operon (transcriptional unit for multiple molecules) and this larger molecule undergoes both tailoring and chemical modifications before each rRNA molecule assumes its mature form. The whole process of biosynthesis of 70S ribosomes takes place in cytoplasm. The required amount of ribonucleoprotein synthesis is under autogenous regulation of translation.

In eukaryotes, the biogenesis of ribosomes is much more complex involving three main events: rRNA synthesis by nucleolar organizer (NO), synthesis of 5S rRNA, and biosynthesis of ribonucleoprotein. The 5.8S, 18S, and 28S rRNAs are transcribed as a much larger molecule in the nucleolar organizer. 5S rRNA is synthesized outside of nucleolus. The ribonucleoproteins are synthesized in the cytoplasm by usual mechanism. It is in the nucleolus that newly synthesized rRNAs accumulate and become associated with required ribonucleoproteins and then migrate to the cytoplasm of cell in the form of ribosomal subunits.

Function

Ribosomes are the workhorses of protein biosynthesis, the process of translating messenger RNA (mRNA) into protein. The mRNA comprises a series of codons that dictate to the ribosome the sequence of the amino acids needed to make the protein. Using the mRNA as a template, the ribosome traverses each codon of the mRNA, pairing it with the appropriate amino acid. This is done using molecules of transfer RNA (tRNA) containing a complementary anticodon on one end and the appropriate amino acid on the another.

Protein synthesis begins at a start codon near the 5' end of the mRNA. The small ribosomal subunit, typically bound to a tRNA containing the amino acid methionine, binds to an AUG codon on the mRNA and recruits the large ribosomal subunit. Of the three tRNA binding sites, designated by A, P, and E, the A–site tRNA bears an incoming amino acid, and the P–site tRNA carries the growing peptide chain. Peptide bond formation attaches the peptide to the A–site tRNA's amino acid. The P–site tRNA then moves to the E–site (E stands for "exit"), replacing the former, uncharged E–site tRNA. The A–site tRNA, now bearing the growing peptide, is shifted into the P position. A new tRNA bearing the next amino acid is then brought into the A–site.

Figure 4 : Translation of mRNA (1) by a ribosome (2) into a polypeptide chain (3). The mRNA begins with a start codon (AUG) and ends with a stop codon (UAG).

In Figure 4, both ribosomal subunits (small and large) assemble at the start codon (towards the 5' end of the mRNA). The ribosome uses tRNA which matches the current codon (triplet) on the mRNA to append an amino acid to the polypeptide chain. This is done for each triplet on the mRNA, while the ribosome moves towards the 3' end of the mRNA. Usually in bacterial cells, several ribosomes are working parallel on a single mRNA, forming what we call a polyribosome or polysome.

References

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• Ban N., Nissen P., Hansen J., Moore P.B., Steitz T.A. 2000. The complete atomic structure of the large ribosomal subunit at 2.4 Å resolution. Science. 289(5481):905–20. PMID 10937989
• Bruce Alberts, Alexander Johnson, Julian Lewis, Martin Raff, Keith Roberts, Peter Walter. 2002. The Molecular Biology of the Cell. Garland Science. Fourth Edition. ISBN 0815332181
• O'Brien, T.W. 1971. The General Occurrence of 55S Ribosomes in Mammalian Liver Mitochondria. J. Biol. Chem. Vol. 245:3409.
• Palade, G.E. 1955. A small particulate component of the cytoplasm. J. Biophys. Biochem. Cytol. Vol. 1(1):59–68. PMID 14381428
• Roberts, R. B. 1958. Introduction in "Microsomal Particles and Protein Synthesis". Pergamon Press, Inc. New York.
• Schluenzen F., Tocilj A., Zarivach R., Harms J., Gluehmann M., Janell D., Bashan A., Bartels H., Agmon I., Franceschi F., Yonath A. 2000. Structure of functionally activated small ribosomal subunit at 3.3 angstroms resolution. Cell. 102(5):615–23. PMID 11007480
• Schuwirth B.S., Borovinskaya M.A., Hau C.W., Zhang W., Vila–Sanjurjo A., Holton J.M., Cate J.H. 2005. Structures of the bacterial ribosome at 3.5 Å resolution. Science. 310(5749):827–34. PMID 16272117
• Selmer, M., Dunham, C.M., Murphy, F.V IV, Weixlbaumer, A., Petry S., Kelley, A.C., Weir, J.R. and Ramakrishnan, V. 2006. Structure of the 70S ribosome complexed with mRNA and tRNA. Science. 313(5795):1935–1942. PMID 16959973
• Wimberly BT, Brodersen DE, Clemons WM Jr, Morgan–Warren RJ, Carter AP, Vonrhein C, Hartsch T, Ramakrishnan V. 2000. Structure of the 30S ribosomal subunit. Nature. 407(6802):327–39. PMID 11014182
• Yusupov M.M., Yusupova G.Z., Baucom A., Lieberman K., Earnest T.N., Cate J.H., Noller H.F. 2001. Crystal structure of the ribosome at 5.5 Å resolution. Science. 292(5518):883–96. PMID 11283358

External links

• Lab computer simulates ribosome in motionRetrieved August 27, 2007.
• Role of ribosomesRetrieved August 27, 2007.
• RCSB Protein Data Bank: RibosomeRetrieved August 27, 2007.
• RCSB Protein Data Bank: Elongation FactorsRetrieved August 27, 2007.

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