Ribosome biogenesis is the process of making ribosomes. This is an energy consuming, dynamic process, requiring the synthesis of around 200 proteins in the processing of ribosomal RNAs and assembling them with ribosomal proteins to make the ribosomes subunits.
The sequence ofDNA that encodes the sequence of theamino acids in a protein is transcribed into amessenger RNA (mRNA) chain. Ribosomes bind to the messenger RNA molecules and use the RNA's sequence ofnucleotides to determine the sequence of amino acids needed to generate a protein. Amino acids are selected and carried to the ribosome bytransfer RNA (tRNA) molecules, which enter the ribosome and bind to the messenger RNA chain via ananticodon stem loop. For each coding triplet (codon) in the messenger RNA, there is a unique transfer RNA that must have the exact anti-codon match, and carries the correct amino acid for incorporating into a growingpolypeptide chain. Once the protein is produced, it can thenfold to produce a functionalthree-dimensional structure.[1][2]
During translation the synthesis of proteins from their building blocks takes place in four stages: initiation, elongation, termination, and ribosome recycling.[3] Thestart codon in all mRNA molecules has the sequence AUG. The stop codon is one of UAA, UAG, or UGA; since there are no tRNA molecules that recognize these codons, the ribosome recognizes that translation is complete.[4] When a ribosome finishes reading an mRNA molecule, the two subunits separate and are usually broken up but can be reused. Ribosomes are a kind ofenzyme, calledribozymes because thecatalyticpeptidyl transferase activity that links amino acids together is performed by the ribosomal RNA.[5]
Ribosomes frombacteria,archaea, andeukaryotes (in thethree-domain system) resemble each other to a remarkable degree, evidence of a common origin. They differ in their size, sequence, structure, and the ratio of protein to RNA. The differences in structure allow someantibiotics to kill bacteria by inhibiting their ribosomes while leaving human ribosomes unaffected. In all domains, apolysome of two or more ribosomes may move along a single mRNA chain at one time, each reading a specific sequence and producing a corresponding protein molecule.[citation needed]
Themitochondrial ribosomes (mitoribosomes) of eukaryotic cells are distinct from the other ribosomes. They functionally resemble those in bacteria, reflecting the evolutionary origin of mitochondria asendosymbiotic bacteria.[6][7]
Ribosomes were first observed in the mid-1950s as dense particles or granules byRomanian-American cell biologistGeorge Emil Palade, using anelectron microscope.[8] They were initially calledPalade granules due to their granular structure.[citation needed] The term "ribosome" was proposed in 1958 by Howard M. Dintzis:[9]
During the course of the symposium a semantic difficulty became apparent. To some of the participants, "microsomes" mean the ribonucleoprotein particles of the microsome fraction contaminated by other protein and lipid material; to others, the microsomes consist of protein and lipid contaminated by particles. The phrase "microsomal particles" does not seem adequate, and "ribonucleoprotein particles of the microsome fraction" is much too awkward. During the meeting, the word "ribosome" was suggested, which has a very satisfactory name and a pleasant sound. The present confusion would be eliminated if "ribosome" were adopted to designate ribonucleoprotein particles in sizes ranging from 35 to 100S.
— Albert Claude, Microsomal Particles and Protein Synthesis[10]
A ribosome is largely made up of specializednon-codingribosomal RNA (rRNA) as well as dozens of distinctribosomal proteins (the number varies slightly between species). The ribosomal proteins and rRNAs are arranged into two distinct ribosomal subunits one large and one small. The subunits fit together locking around a strand of mRNA, and work as one to translate the mRNA into apolypeptide chain duringprotein synthesis.[14]
Bacterial ribosomes are around 20 nm (200 Å) in diameter and are composed of 65% rRNA and 35%ribosomal proteins.[15] Eukaryotic ribosomes are between 25 and 30nm (250–300 Å) in diameter with an rRNA-to-protein ratio that is close to 1.[16]Crystallographic work[17] has shown that there are no ribosomal proteins close to the reaction site for polypeptide synthesis. This suggests that the protein components of ribosomes do not directly participate in peptide bond formation catalysis, but rather that these proteins act as a scaffold that may enhance the ability of rRNA to synthesize protein[18]
Molecular structure of the 30S subunit fromThermus thermophilus.[19] Proteins are shown in blue and the single RNA chain in brown.
The unit of measurement used to describe the ribosomal subunits and the rRNA fragments is theSvedberg unit, a measure of the rate ofsedimentation incentrifugation rather than size. This accounts for why fragment names do not add up: for example, bacterial 70S ribosomes are made of 50S and 30S subunits.[citation needed]
Prokaryotes have 70S ribosomes, each consisting of a small (30S) and a large (50S) subunit.E. coli, for example, has a16S RNA subunit (consisting of 1540 nucleotides) that is bound to 21 proteins. The large subunit is composed of a5S RNA subunit (120 nucleotides), a 23S RNA subunit (2900 nucleotides) and 31proteins.[13]
Affinity label for the tRNA binding sites on theE. coli ribosome allowed the identification of A and P site proteins most likely associated with the peptidyltransferase activity;[5] labelled proteins are L27, L14, L15, L16, L2; at least L27 is located at the donor site, as shown by E. Collatz and A.P. Czernilofsky.[21][22] Additional research has demonstrated that the S1 and S21 proteins, in association with the 3′-end of 16S ribosomal RNA, are involved in the initiation of translation.[23]
Archaeal ribosomes are conventionally quoted as having similar sizes as the bacterial ribosome, being a 70S ribosome made up from a 50S large subunit and a 30S small subunit.[24] The rRNA chains are similarly commonly called 16S, 23S, and 5S, though again few (if any) recent sources have truly measured theirsedimentation coefficients.[25] However, on the sequence and structual levels, they are much closer to eukaryotic ones than to bacterial ones. Every extra ribosomal protein archaea have compared to bacteria has a eukaryotic counterpart, while no such relation applies between archaea and bacteria.[26][27][28]
During 1977, Czernilofsky published research that usedaffinity labeling to identify tRNA-binding sites on rat liver ribosomes. Several proteins, including L32/33, L36, L21, L23, L28/29 and L13 were implicated as being at or near thepeptidyl transferase center.[34]
In eukaryotes, ribosomes are present inmitochondria (sometimes calledmitoribosomes) and inplastids such aschloroplasts (also called plastoribosomes). They also consist of large and small subunits bound together withproteins into one 70S particle.[13] These ribosomes are similar to those of bacteria and these organelles are thought to have originated assymbiotic bacteria.[13] Of the two, chloroplastic ribosomes are closer to bacterial ones than mitochondrial ones are. Many pieces of ribosomal RNA in the mitochondria are shortened, and in the case of5S rRNA, replaced by other structures in animals and fungi.[35] In particular,Leishmania tarentolae has a minimalized set of mitochondrial rRNA.[36] In contrast, plant mitoribosomes have both extended rRNA and additional proteins as compared to bacteria, in particular, many pentatricopetide repeat proteins.[37]
Thecryptomonad andchlorarachniophyte algae may contain anucleomorph that resembles a vestigial eukaryotic nucleus.[38] Eukaryotic 80S ribosomes may be present in the compartment containing the nucleomorph.[39]
The differences between the bacterial and eukaryotic ribosomes are exploited bypharmaceutical chemists to createantibiotics 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 while the eukaryotic 80S ribosomes are not.[40] Even thoughmitochondria 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 theorganelle.[41] A noteworthy counterexample is the antineoplastic antibioticchloramphenicol, which inhibits bacterial 50S and eukaryotic mitochondrial 50S ribosomes.[42] Ribosomes in chloroplasts, however, are different: Antibiotic resistance in chloroplast ribosomal proteins is a trait that has to be introduced as a marker, with genetic engineering.[43]
The various ribosomes share a core structure, which is quite similar despite the large differences in size. Much of the RNA is highly organized into varioustertiary structural motifs, for examplepseudoknots that exhibitcoaxial stacking. The extraRNA in the larger ribosomes is in several long continuous insertions,[44] such that they form loops out of the core structure without disrupting or changing it.[13] All of the catalytic activity of the ribosome is carried out by theRNA; the proteins reside on the surface and seem to stabilize the structure.[13]
Figure 4: Atomic structure of the 50S subunit fromHaloarcula marismortui. Proteins are shown in blue and the two RNA chains in brown and yellow.[45] The small patch of green in the center of the subunit is the active site.
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, of the order of a fewångströms.[46]
The first papers giving the structure of the ribosome at atomic resolution were published almost simultaneously in late 2000. The 50S (large prokaryotic) subunit was determined from thearchaeonHaloarcula marismortui[45] and thebacteriumDeinococcus radiodurans, and the structure of the 30S subunit was determined from the bacteriumThermus thermophilus.[19][47] These structural studies were awarded the Nobel Prize in Chemistry in 2009. In May 2001 these coordinates were used to reconstruct the entireT. thermophilus 70S particle at 5.5 Å resolution.[48]
Two papers were published in November 2005 with structures of theEscherichia coli 70S ribosome. The structures of a vacant ribosome were determined at 3.5 Å resolution usingX-ray crystallography.[49] Then, two weeks later, a structure based oncryo-electron microscopy was published,[50] which depicts the ribosome at 11–15 Å resolution in the act of passing a newly synthesized protein strand into the protein-conducting channel.
The first atomic structures of the ribosome complexed withtRNA andmRNA molecules were solved by using X-ray crystallography by two groups independently, at 2.8 Å[51] and at 3.7 Å.[52] These structures allow one to see the details of interactions of theThermus thermophilus ribosome withmRNA and withtRNAs bound at classical ribosomal sites. Interactions of the ribosome with long mRNAs containingShine-Dalgarno sequences were visualized soon after that at 4.5–5.5 Å resolution.[53] In 2023, a cryo-electron microscopy study reported a 1.55 Å structure of theEscherichia coli 70S ribosome in the translating state, providing near-atomic detail of rRNA modifications, tRNA-mRNA interactions, and ion coordination. The high-resolution map enabled identification of ribosomal polymorphism sites and visualization of transient chimeric hybrid states associated with tRNA translocation at approximately 2 Å resolution. These findings improved structural understanding of the ribosome's functional regions and offered valuable insights for antibiotic design.[54]
In 2011, the first complete atomic structure of the eukaryotic 80S ribosome from the yeastSaccharomyces cerevisiae was obtained by crystallography.[31] The model reveals the architecture of eukaryote-specific elements and their interaction with the universally conserved core. At the same time, the complete model of a eukaryotic 40S ribosomal structure inTetrahymena thermophila was published and described the structure of the40S subunit, as well as much about the 40S subunit's interaction witheIF1 duringtranslation initiation.[32] Similarly, the eukaryotic60S subunit structure was also determined fromTetrahymena thermophila in complex witheIF6.[33] In addition, high-resolution cryo-EM structures of a thermophilic eukaryotic 80S ribosome captured in two rotational states at ~2.9 Å and ~3.0 Å resolution revealed atomistic details of the eukaryotic translocation mechanism and conformational dynamics of eEF2 during GTP hydrolysis.[55]
Duringtranslation, tRNA charged with amino acid enters the ribosome and aligns with the correct mRNA triplet. Ribosome then adds amino acid to growing protein chain.
Ribosomes are minute particles consisting of RNA and associated proteins that function to synthesize proteins. Proteins are needed for many cellular functions, such as repairing damage or directing chemical processes. Ribosomes can be either free, floating within the cytoplasm or attached to therough endoplasmic reticulum. Their main function is to convert genetic code into an amino acid sequence and to build protein polymers from amino acid monomers.[citation needed]
Ribosomes act as catalysts in two extremely important biological processes called peptidyl transfer and peptidyl hydrolysis.[5][56] The "PT center is responsible for producing protein bonds during protein elongation".[56]
In summary, ribosomes have two main functions: Decoding the message, and the formation of peptide bonds. These two functions reside in the ribosomal subunits. Each subunit is made of one or more rRNAs and many r-proteins. The small subunit (30S in bacteria and archaea, 40S in eukaryotes) has the decoding function, whereas the large subunit (50S in bacteria and archaea, 60S in eukaryotes) catalyzes the formation of peptide bonds, referred to as the peptidyl-transferase activity. The bacterial (and archaeal) small subunit contains the 16S rRNA and 21 r-proteins (Escherichia coli), whereas the eukaryotic small subunit contains the 18S rRNA and 32 r-proteins (Saccharomyces cerevisiae, although the numbers vary between species). The bacterial large subunit contains the 5S and 23S rRNAs and 34 r-proteins (E. coli), with the eukaryotic large subunit containing the 5S, 5.8S, and 25S/28S rRNAs and 46 r-proteins (S. cerevisiae; again, the exact numbers vary between species).[57]
Ribosomes are the workplaces ofprotein biosynthesis, the process of translatingmRNA intoprotein. The mRNA comprises a series ofcodons which are decoded by the ribosome to make the protein. Using the mRNA as a template, the ribosome traverses each codon (3 nucleotides) of the mRNA, pairing it with the appropriate amino acid provided by anaminoacyl-tRNA. Aminoacyl-tRNA contains a complementaryanticodon on one end and the appropriate amino acid on the other. For fast and accurate recognition of the appropriate tRNA, the ribosome utilizes large conformational changes (conformational proofreading).[58] The small ribosomal subunit, typically bound to an aminoacyl-tRNA containing the first amino acidmethionine, binds to an AUG codon on the mRNA and recruits the large ribosomal subunit. The ribosome contains three RNA binding sites, designated A, P, and E. TheA-site binds an aminoacyl-tRNA or termination release factors;[59][60] theP-site binds a peptidyl-tRNA (a tRNA bound to the poly-peptide chain); and theE-site (exit) binds a free tRNA. Protein synthesis begins at astart codon AUG near the5′-end of the mRNA. mRNA binds to the P site of the ribosome first. The ribosome recognizes the start codon by using theShine-Dalgarno sequence of the mRNA in prokaryotes andKozak box in eukaryotes.[citation needed]
Although catalysis of thepeptide bond involves the C2hydroxyl of RNA's P-siteadenosine in a proton shuttle mechanism, other steps in protein synthesis (such as translocation) are caused by changes in protein conformations. Since theircatalytic core is made of RNA, ribosomes are classified as "ribozymes,"[61] and it is thought that they might be remnants of theRNA world.[62]
Figure 5: Translation of mRNA (1) by a ribosome (2)(shown assmall andlarge subunits) into apolypeptide chain (3). The ribosome begins at the start codon of RNA (AUG) and ends at the stop codon (UAG).
In Figure 5, both ribosomal subunits (small andlarge) assemble at the start codon (towards the 5' end of themRNA). The ribosome usestRNA that matches the current codon (triplet) on the mRNA to append anamino 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 is called apolyribosome orpolysome.[citation needed]
The ribosome is known to actively participate in theprotein folding.[63][64] The structures obtained in this way are usually identical to the ones obtained during protein chemical refolding; however, the pathways leading to the final product may be different.[65][66] In some cases, the ribosome is crucial in obtaining the functional protein form. For example, one of the possible mechanisms of folding of the deeplyknotted proteins relies on the ribosome pushing the chain through the attached loop.[67]
Presence of a ribosome quality control protein Rqc2 is associated with mRNA-independent protein elongation.[68][69] This elongation is a result of ribosomal addition (via tRNAs brought by Rqc2) ofCAT tails: ribosomes extend theC-terminus of a stalled protein with random, translation-independent sequences ofalanines andthreonines.[70][71]
Ribosomes are classified as being either "free" or "membrane-bound".[72]Free and membrane-bound ribosomes differ only in their spatial distribution; they are identical in structure. Whether the ribosome exists in a free or membrane-bound state depends on the presence of anER-targeting signal sequence on the protein being synthesized, so an individual ribosome might be membrane-bound when it is making one protein, but free in the cytosol when it makes another protein.
Ribosomes are sometimes referred to asorganelles, but the use of the termorganelle is often restricted to describing sub-cellular components that include a phospholipid membrane, which ribosomes, being entirely particulate, do not. For this reason, ribosomes may sometimes be described as "non-membranous organelles".[citation needed]
Free ribosomes can move about anywhere in thecytosol, but are excluded from thecell nucleus and other organelles. Proteins that are formed from free ribosomes are released into the cytosol and used within the cell. Since the cytosol contains high concentrations ofglutathione and is, therefore, areducing environment, proteins containingdisulfide bonds, which are formed from oxidized cysteine residues, cannot be produced within it.[citation needed]
When a ribosome begins to synthesize proteins needed in certain organelles, the ribosome making this protein can become "membrane-bound". In eukaryotic cells this happens in a region of the endoplasmic reticulum (ER) called the "rough ER". The newly produced polypeptide chains are inserted directly into the ER by the ribosome undertakingvectorial synthesis and are then transported to their destinations, through thesecretory pathway. Bound ribosomes usually produce proteins that are used within the plasma membrane or are expelled from the cell viaexocytosis.[73]
In bacterial cells, ribosomes are synthesized in the cytoplasm through thetranscription of multiple ribosome geneoperons. In eukaryotes, the process takes place both in the cell cytoplasm and in thenucleolus, which is a region within thecell nucleus. The assembly process involves the coordinated function of over 200 proteins in the synthesis and processing of the four rRNAs, as well as assembly of those rRNAs with the ribosomal proteins.[74]
The ribosome may have first originated as a protoribosome,[75] possibly containing a peptidyl transferase centre (PTC), in anRNA world, appearing as a self-replicating complex that only later evolved the ability to synthesize proteins whenamino acids began to appear.[76] Studies suggest that ancient ribosomes constructed solely ofrRNA could have developed the ability to synthesizepeptide bonds.[77][78][79][80][81] In addition, evidence strongly points to ancient ribosomes as self-replicating complexes, where the rRNA in the ribosomes had informational, structural, and catalytic purposes because it could have coded fortRNAs and proteins needed for ribosomal self-replication.[82] Hypothetical cellular organisms with self-replicating RNA but without DNA are calledribocytes (or ribocells).[83][84]
As amino acids gradually appeared in the RNA world under prebiotic conditions,[85][86] their interactions with catalytic RNA would increase both the range and efficiency of function of catalytic RNA molecules.[76] Thus, the driving force for the evolution of the ribosome from an ancientself-replicating machine into its current form as a translational machine may have been the selective pressure to incorporate proteins into the ribosome's self-replicating mechanisms, so as to increase its capacity for self-replication.[82][87][88]
In 1958, Francis Crick famously proposed the "one gene-one ribosome-one protein hypothesis," where each ribosome carries the genetic information required to encode a single protein. Although discredited at that time, from the discovery of the first ribosomopathy Diamond–Blackfan Anemia in 1999, the ribosome has transitioned from a passive molecular machine to a dynamic macromolecular machine[89][90].Ribosomes are compositionally heterogeneous between species and even within the same cell, as evidenced by the existence of cytoplasmic and mitochondria ribosomes within the same eukaryotic cells. Certain researchers have suggested that heterogeneity in the composition of ribosomal proteins in mammals is important for gene regulation,i.e., the specialized ribosome hypothesis.[91][92] However, this hypothesis is controversial and the topic of ongoing research.[93][94]
Heterogeneity in ribosome composition was first proposed to be involved in translational control of protein synthesis by Vince Mauro andGerald Edelman.[95] They proposed the ribosome filter hypothesis to explain the regulatory functions of ribosomes. Evidence has suggested that specialized ribosomes specific to different cell populations may affect how genes are translated.[96] Some ribosomal proteins exchange from the assembled complex withcytosolic copies[97] suggesting that the structure of thein vivo ribosome can be modified without synthesizing an entire new ribosome.
Certain ribosomal proteins are absolutely critical for cellular life while others are not. Inbudding yeast, 14/78 ribosomal proteins are non-essential for growth, while in humans this depends on the cell of study.[98] Other forms of heterogeneity include post-translational modifications to ribosomal proteins such as acetylation, methylation, and phosphorylation.[99]Arabidopsis,[100][101][102][103] Viralinternal ribosome entry sites (IRESs) may mediate translations by compositionally distinct ribosomes. For example, 40S ribosomal units withouteS25 in yeast and mammalian cells are unable to recruit theCrPV IGR IRES.[104]
Heterogeneity of ribosomal RNA modifications plays a significant role in structural maintenance and/or function and most mRNA modifications are found in highly conserved regions.[105][106] The most common rRNA modifications arepseudouridylation and2'-O-methylation ofribose.[107]
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