Rules by which information encoded within genetic material is translated into proteins
A series of codons in part of amessenger RNA (mRNA) molecule. Each codon consists of threenucleotides, usually corresponding to a singleamino acid. The nucleotides are abbreviated with the letters A, U, G and C. This is mRNA, which uses U (uracil). DNA uses T (thymine) instead. This mRNA molecule will instruct aribosome to synthesize a protein according to this code.
Genetic code is a set of rules used by livingcells totranslate information encoded within genetic material (DNA orRNA sequences of nucleotide triplets orcodons) intoproteins. Translation is accomplished by theribosome, which linksproteinogenic amino acids in an order specified bymessenger RNA (mRNA), usingtransfer RNA (tRNA) molecules to carry amino acids and to read the mRNA threenucleotides at a time. The genetic code is highly similar among all organisms and can be expressed in a simple table with 64 entries.
The codons specify which amino acid will be added next duringprotein biosynthesis. With some exceptions,[1] a three-nucleotide codon in a nucleic acid sequence specifies a single amino acid. The vast majority ofgenes are encoded with a single scheme (see theRNA codon table). That scheme is often called the canonical or standard genetic code, or simplythe genetic code, thoughvariant codes (such as inmitochondria) exist.
Efforts to understand how proteins are encoded began afterDNA's structure was discovered in 1953. The key discoverers, English biophysicistFrancis Crick and American biologistJames Watson, working together at theCavendish Laboratory of the University of Cambridge, hypothesied that information flows from DNA and that there is a link between DNA and proteins.[2] Soviet-American physicistGeorge Gamow was the first to give a workable scheme for protein synthesis from DNA.[3] He postulated that sets of three bases (triplets) must be employed to encode the 20 standard amino acids used by living cells to build proteins, which would allow a maximum of4‹ThetemplateSmallsup is beingconsidered for deletion.›3 = 64 amino acids (all permutations of the four bases, read three at a time).[4] He named this DNA–protein interaction (the original genetic code) as the "diamond code".[5]
In 1954, Gamow created an informal scientific organisation theRNA Tie Club, as suggested by Watson, for scientists of different persuasions who were interested in howproteins were synthesised from genes. However, the club could have only 20 permanent members to represent each of the 20 amino acids; and four additional honorary members to represent the four nucleotides of DNA.[6]
The first scientific contribution of the club, later recorded as "one of the most important unpublished articles in the history of science"[7] and "the most famous unpublished paper in the annals of molecular biology",[8] was made by Crick. Crick presented a type-written paper titled "On Degenerate Templates and the Adaptor Hypothesis: A Note for the RNA Tie Club"[9] to the members of the club in January 1955, which "totally changed the way we thought about protein synthesis", as Watson recalled.[10] The hypothesis states that the triplet code was not passed on to amino acids as Gamow thought, but carried by a different molecule, an adaptor, that interacts with amino acids.[8] The adaptor was later identified as tRNA.[11]
This was followed by experiments inSevero Ochoa's laboratory that demonstrated that the poly-adenine RNA sequence (AAAAA...) coded for the polypeptide poly-lysine[14] and that the poly-cytosine RNA sequence (CCCCC...) coded for the polypeptide poly-proline.[15] Therefore, the codon AAA specified the amino acidlysine, and the codon CCC specified the amino acidproline. Using variouscopolymers most of the remaining codons were then determined.
Subsequent work byHar Gobind Khorana identified the rest of the genetic code. Shortly thereafter,Robert W. Holley determined the structure oftransfer RNA (tRNA), the adapter molecule that facilitates the process of translating RNA into protein. This work was based upon Ochoa's earlier studies, yielding the latter theNobel Prize in Physiology or Medicine in 1959 for work on theenzymology of RNA synthesis.[16]
Extending this work, Nirenberg andPhilip Leder revealed the code's triplet nature and deciphered its codons. In these experiments, various combinations ofmRNA were passed through a filter that containedribosomes, the components of cells thattranslate RNA into protein. Unique triplets promoted the binding of specific tRNAs to the ribosome. Leder and Nirenberg were able to determine the sequences of 54 out of 64 codons in their experiments.[17] Khorana, Holley and Nirenberg received the Nobel Prize (1968) for their work.[18]
The three stop codons were named by discoverers Richard Epstein and Charles Steinberg. "Amber" was named after their friend Harris Bernstein, whose last name means "amber" in German.[19] The other two stop codons were named "ochre" and "opal" in order to keep the "color names" theme.
In a broad academic audience, the concept of the evolution of the genetic code from the original and ambiguous genetic code to a well-defined ("frozen") code with the repertoire of 20 (+2) canonical amino acids is widely accepted.[20]However, there are different opinions, concepts, approaches and ideas, which is the best way to change it experimentally.[clarification needed] Even models are proposed that predict "entry points" for synthetic amino acid invasion of the genetic code.[21]
Since 2001, 40 non-natural amino acids have been added into proteins by creating a unique codon (recoding) and a corresponding transfer-RNA:aminoacyl – tRNA-synthetase pair to encode it with diverse physicochemical and biological properties in order to be used as a tool to exploringprotein structure and function or to create novel or enhanced proteins.[22][23]
H. Murakami and M. Sisido extended some codons to have four and five bases.Steven A. Benner constructed a functional 65th (in vivo) codon.[24]
In 2015N. Budisa,D. Söll and co-workers reported the full substitution of all 20,899tryptophan residues (UGG codons) with unnatural thienopyrrole-alanine in the genetic code of thebacteriumE. coli.[25]
In 2016 the first stable semisynthetic organism was created. It was a (single cell) bacterium with two synthetic bases (called X and Y). The bases survived cell division.[26][27]
In 2017, researchers in South Korea reported that they had engineered a mouse with an extended genetic code that can produce proteins with unnatural amino acids.[28]
In May 2019, researchers reported the creation of a new "Syn61" strain of theE. coli bacteria. This strain has a fullysynthetic genome that is refactored (all overlaps expanded), recoded (removing the use of three out of 64 codons completely), and further modified to remove the now unnecessary tRNAs and release factors. It is fullyviable and grows 1.6× slower than its wild-type counterpart "MDS42".[29][30]
In 2025, researchers reported a new "Syn57" strain, which removes the use of 7 out of 64 codons completely.[31]
Reading frames in the DNA sequence of a region of the human mitochondrial genome coding for the genesMT-ATP8 andMT-ATP6 (in black: positions 8,525 to 8,580 in the sequence accession NC_012920[32]). There are three possible reading frames in the 5' → 3' forward direction, starting on the first (+1), second (+2) and third position (+3). For each codon (square brackets), the amino acid is given by thevertebrate mitochondrial code, either in the +1 frame forMT-ATP8 (in red) or in the +3 frame forMT-ATP6 (in blue). TheMT-ATP8 genes terminates with the TAG stop codon (red dot) in the +1 frame. TheMT-ATP6 gene starts with the ATG codon (blue circle for the M amino acid) in the +3 frame.
A reading frame is defined by the initial triplet of nucleotides from which translation starts. It sets the frame for a run of successive, non-overlapping codons, which is known as an "open reading frame" (ORF). For example, the string 5'-AAATGAACG-3' (see figure), if read from the first position, contains the codons AAA, TGA, and ACG ; if read from the second position, it contains the codons AAT and GAA ; and if read from the third position, it contains the codons ATG and AAC. Every sequence can, thus, be read in its5' → 3' direction in threereading frames, each producing a possibly distinct amino acid sequence: in the given example, Lys (K)-Trp (W)-Thr (T), Asn (N)-Glu (E), or Met (M)-Asn (N), respectively (when translating with thevertebrate mitochondrial code). When DNA is double-stranded, six possiblereading frames are defined, three in the forward orientation on one strand and three reverse on the opposite strand.[33]: 330 Protein-coding frames are defined by astart codon, usually the first AUG codon in the RNA, (ATG in DNA) sequence.
Translation starts with a chain-initiation codon orstart codon. The start codon alone is not sufficient to begin the process. Nearby sequences such as theShine-Dalgarno sequence inE. coli andinitiation factors are also required to start translation. The most common start codon is AUG, which is read asmethionine or asformylmethionine (in bacteria, mitochondria, and plastids). Alternative start codons depending on the organism include "GUG" or "UUG"; these codons normally representvaline andleucine, respectively, but as start codons they are translated as methionine or formylmethionine.[34]
The threestop codons have names: UAG isamber, UGA isopal (sometimes also calledumber), and UAA isochre. Stop codons are also called "termination" or "nonsense" codons. They signal release of the nascent polypeptide from the ribosome because no cognate tRNA has anticodons complementary to these stop signals, allowing arelease factor to bind to the ribosome instead.[35]
Examples of notablemutations that can occur in humans[36]
During the process ofDNA replication, errors occasionally occur in thepolymerization of the second strand. These errors,mutations, can affect an organism'sphenotype, especially if they occur within the protein coding sequence of a gene. Error rates are typically 1 error in every 10–100 million bases—due to the "proofreading" ability ofDNA polymerases.[37][38]
Mutations that disrupt the reading frame sequence byindels (insertions ordeletions) of a non-multiple of 3 nucleotide bases are known asframeshift mutations. These mutations usually result in a completely different translation from the original, and likely cause astop codon to be read, which truncates the protein.[41] These mutations may impair the protein's function and are thus rare inin vivo protein-coding sequences. One reason inheritance of frameshift mutations is rare is that, if the protein being translated is essential for growth under the selective pressures the organism faces, absence of a functional protein may cause death before the organism becomes viable.[42] Frameshift mutations may result in severe genetic diseases such asTay–Sachs disease.[43]
Although most mutations that change protein sequences are harmful or neutral, some mutations have benefits.[44] These mutations may enable the mutant organism to withstand particular environmental stresses better thanwild type organisms, or reproduce more quickly. In these cases a mutation will tend to become more common in a population throughnatural selection.[45]Viruses that useRNA as their genetic material have rapid mutation rates,[46] which can be an advantage, since these viruses thereby evolve rapidly, and thus evade theimmune system defensive responses.[47] In large populations of asexually reproducing organisms, for example,E. coli, multiple beneficial mutations may co-occur. This phenomenon is calledclonal interference and causes competition among the mutations.[48]
Grouping of codons by amino acid residue molar volume andhydropathicity. Amore detailed version is available.Axes 1, 2, 3 are the first, second, and third positions in the codon. The 20 amino acids and stop codons (X) are shown insingle letter code.
Degeneracy is the redundancy of the genetic code. This term was given by Bernfield and Nirenberg. The genetic code has redundancy but no ambiguity (see thecodon tables below for the full correlation). For example, although codons GAA and GAG both specifyglutamic acid (redundancy), neither specifies another amino acid (no ambiguity). The codons encoding one amino acid may differ in any of their three positions. For example, the amino acid leucine is specified byYUR or CUN (UUA, UUG, CUU, CUC, CUA, or CUG) codons (difference in the first or third position indicated usingIUPAC notation), while the amino acidserine is specified by UCN or AGY (UCA, UCG, UCC, UCU, AGU, or AGC) codons (difference in the first, second, or third position).[49] A practical consequence of redundancy is that errors in the third position of the triplet codon cause only a silent mutation or an error that would not affect the protein because thehydrophilicity orhydrophobicity is maintained by equivalent substitution of amino acids; for example, a codon of NUN (where N = any nucleotide) tends to code for hydrophobic amino acids. NCN yields amino acid residues that are small in size and moderate inhydropathicity; NAN encodes average size hydrophilic residues. The genetic code is so well-structured for hydropathicity that a mathematical analysis (Singular Value Decomposition) of 12 variables (4 nucleotides x 3 positions) yields a remarkable correlation (C = 0.95) for predicting the hydropathicity of the encoded amino acid directly from the triplet nucleotide sequence,without translation.[50][51] Note in the table, below, eight amino acids are not affected at all by mutations at the third position of the codon, whereas in the figure above, a mutation at the second position is likely to cause a radical change in the physicochemical properties of the encoded amino acid.Nevertheless, changes in the first position of the codons are more important than changes in the second position on a global scale.[52] The reason may be that charge reversal (from a positive to a negative charge or vice versa) can only occur upon mutations in the first position of certain codons, but not upon changes in the second position of any codon. Such charge reversal may have dramatic consequences for the structure or function of a protein. This aspect may have been largely underestimated by previous studies.[52]
The frequency of codons, also known ascodon usage bias, can vary from species to species with functional implications for the control oftranslation. The codon varies by organism; for example, most common proline codon in E. coli is CCG, whereas in humans this is the least used proline codon.[53]
In some proteins, non-standard amino acids are substituted for standard stop codons, depending on associated signal sequences in the messenger RNA. For example, UGA can code forselenocysteine and UAG can code forpyrrolysine. Selenocysteine came to be seen as the 21st amino acid, and pyrrolysine as the 22nd.[55] Both selenocysteine and pyrrolysine may be present in the same organism.[55] Although the genetic code is normally fixed in an organism, the achaeal prokaryoteAcetohalobium arabaticum can expand its genetic code from 20 to 21 amino acids (by including pyrrolysine) under different conditions of growth.[56]
Genetic codelogo of theGlobobulimina pseudospinescens mitochondrial genome by FACIL. The program is able to correctly infer that theProtozoan Mitochondrial Code is in use.[57] The logo shows the 64 codons from left to right, predicted alternatives in red (relative to the standard genetic code). Red line: stop codons. The height of each amino acid in the stack shows how often it is aligned to the codon in homologous protein domains. The stack height indicates the support for the prediction.
There was originally a simple and widely accepted argument that the genetic code should be universal: namely, that any variation in the genetic code would be lethal to the organism (although Crick had stated that viruses were an exception). This is known as the "frozen accident" argument for the universality of the genetic code. However, in his seminal paper on the origins of the genetic code in 1968, Francis Crick still stated that the universality of the genetic code in all organisms was an unproven assumption, and was probably not true in some instances. He predicted that "The code is universal (the same in all organisms) or nearly so".[58] The first variation was discovered in 1979, by researchers studyinghuman mitochondrial genes.[59] Many slight variants were discovered thereafter,[60] including various alternative mitochondrial codes.[61] These minor variants for example involve translation of the codon UGA astryptophan inMycoplasma species, and translation of CUG as a serine rather than leucine in yeasts of the "CTG clade" (such asCandida albicans).[62][63][64] Because viruses must use the same genetic code as their hosts, modifications to the standard genetic code could interfere with viral protein synthesis or functioning. However, viruses such astotiviruses have adapted to the host's genetic code modification.[65] Inbacteria andarchaea, GUG and UUG are common start codons. In rare cases, certain proteins may use alternative start codons.[60]Surprisingly, variations in the interpretation of the genetic code exist also in human nuclear-encoded genes: In 2016, researchers studying the translation of malate dehydrogenase found that in about 4% of the mRNAs encoding this enzyme the stop codon is naturally used to encode the amino acids tryptophan and arginine.[66] This type of recoding is induced by a high-readthrough stop codon context[67] and it is referred to asfunctional translational readthrough.[68]
Despite these differences, all known naturally occurring codes are very similar. The coding mechanism is the same for all organisms: three-base codons,tRNA, ribosomes, single direction reading and translating single codons into single amino acids.[69] The most extreme variations occur in certain ciliates where the meaning of stop codons depends on their position within mRNA. When close to the 3' end they act as terminators while in internal positions they either code for amino acids as inCondylostoma magnum[70] or triggerribosomal frameshifting as inEuplotes.[71]
The origins and variation of the genetic code, including the mechanisms behind the evolvability of the genetic code, have been widely studied,[72][73] and some studies have been done experimentally evolving the genetic code of some organisms.[74][75][76]
Variant genetic codes used by an organism can be inferred by identifying highly conserved genes encoded in that genome, and comparing its codon usage to the amino acids in homologous proteins of other organisms. For example, the program FACIL infers a genetic code by searching which amino acids in homologous protein domains are most often aligned to every codon. The resulting amino acid (or stop codon) probabilities for each codon are displayed in a genetic code logo.[57]
As of January 2022, the most complete survey of genetic codes is done by Shulgina and Eddy, who screened 250,000 prokaryotic genomes using their Codetta tool. This tool uses a similar approach to FACIL with a largerPfam database. Despite the NCBI already providing 27 translation tables, the authors were able to find new 5 genetic code variations (corroborated by tRNA mutations) and correct several misattributions.[77] Codetta was later used to analyze genetic code change inciliates.[78]
The genetic code is a key part of thehistory of life. Under theRNA world hypothesis, self-replicating RNA molecules preceded significant use of proteins. Under the nucleopeptide world hypothesis, significant use of peptides preceded the genetic code and was concurrent with early life's sophisticated use of RNA.[79] Transfer RNA molecules appear to have evolved before modernaminoacyl-tRNA synthetases.[80] It is possible that synthetases replaced an earlier system ofribozymes (RNA enzymes), or that amino acids were recognized by unique pockets in the tertiary structure of proto-tRNAs.[81] It is not known why the genetic code only uses L-amino acids and not D-amino acids.[82]
Any evolutionary model for the code's origin must account for itsrobustness of encoded proteins to errors during DNA replication and during translation. Many single nucleotide errors aresynonymous, and those that are not tend to cause thesubstitution of a biochemically similar amino acid. Even holding the structure of the code the same such that clusters of codons encode the same amino acid, which amino acids are encoded by which sets of codons is "one in a million" with respect to robustness.[83] Biochemically similar amino acids tend to share the same middle nucleotide, while synonymous changes generally happen at the third nucleotide.
Amino acids that share the same biosynthetic pathway tend to have the same first base in their codons. This could be an evolutionary relic of an early, simpler genetic code with fewer amino acids that later evolved to code a larger set of amino acids.[84] It could also reflect steric and chemical properties that had another effect on the codon during its evolution. Amino acids with similar physical properties also tend to have similar codons,[85][86] reducing the problems caused by point mutations and mistranslations.[83]
Three main hypotheses address the origin of the genetic code. Many models belong to one of them or to a hybrid:[87]
Random freeze: the genetic code was randomly created. For example, earlytRNA-like ribozymes may have had different affinities for amino acids, with codons emerging from another part of the ribozyme that exhibited random variability. Once enoughpeptides were coded for, any major random change in the genetic code would have been lethal; hence it became "frozen".[58]
Stereochemical affinity: the genetic code is a result of a high affinity between each amino acid and its codon or anti-codon; the latter option implies that pre-tRNA molecules matched their corresponding amino acids by this affinity. Later during evolution, this matching was gradually replaced with matching by aminoacyl-tRNA synthetases.[82][88][89]
Optimality: the genetic code continued to evolve after its initial creation, so that the current code maximizes somefitness function, usually some kind of error minimization.[82][87][90]
Hypotheses have addressed a variety of scenarios:[91]
Chemical principles govern specific RNA interaction with amino acids. Experiments withaptamers showed that some amino acids have a selective chemical affinity for their codons.[92] Experiments showed that of 8 amino acids tested, 6 show some RNA triplet-amino acid association.[93][89]
Biosynthetic expansion. The genetic code grew from a simpler earlier code through a process of "biosynthetic expansion". Primordial life "discovered" new amino acids (for example, as by-products ofmetabolism) and later incorporated some of these into the machinery of genetic coding.[73] Although much circumstantial evidence has been found to suggest that fewer amino acid types were used in the past,[94] precise and detailed hypotheses about which amino acids entered the code in what order are controversial.[95][96] However, several studies have suggested that Gly, Ala, Asp, Val, Ser, Pro, Glu, Leu, Thr may belong to a group of early-addition amino acids, whereas Cys, Met, Tyr, Trp, His, Phe may belong to a group of later-addition amino acids.[97][98][99][100] An alternative analysis of amino acid usage in theLast Universal Common Ancestor concluded that the amino acids came in the following order: Val, Gly, Ile, Met, Ala, Thr, His, Glu, Cys, Pro, Lys, Ser, Asp, Leu, Asn, Arg, Phe, Tyr, Gln, Trp.[101] It was pointed out that the late appearance of sulfur-containing cysteine and methionine was concluded in part from their absence of the sulfur-freeMiller–Urey experiment, that early life is believed to have usedS-adenosyl methionine, and that while histidine is hard to make abiotically, it is straightforward to synthesize in an organism that already has sophisticated RNA and hence purine synthesis.[101]
Natural selection has led to codon assignments of the genetic code that minimize the effects ofmutations.[102] A recent hypothesis[103] suggests that the triplet code was derived from codes that used longer than triplet codons (such as quadruplet codons). Longer than triplet decoding would increase codon redundancy and would be more error resistant. This feature could allow accurate decoding absent complex translational machinery such as theribosome, such as before cells began making ribosomes.
Information channels:Information-theoretic approaches model the process of translating the genetic code into corresponding amino acids as an error-prone information channel.[104] The inherent noise (that is, the error) in the channel poses the organism with a fundamental question: how can a genetic code be constructed to withstand noise[105] while accurately and efficiently translating information? These"rate-distortion" models[106] suggest that the genetic code originated as a result of the interplay of the three conflicting evolutionary forces: the needs for diverse amino acids,[107] for error-tolerance[102] and for minimal resource cost. The code emerges at a transition when the mapping of codons to amino acids becomes nonrandom. The code's emergence is governed by thetopology defined by the probable errors and is related to themap coloring problem.[108]
Game theory: Models based onsignaling games combine elements of game theory, natural selection and information channels. Such models have been used to suggest that the first polypeptides were likely short and had non-enzymatic function. Game theoretic models suggested that the organization of RNA strings into cells may have been necessary to prevent "deceptive" use of the genetic code, i.e. preventing the ancient equivalent of viruses from overwhelming the RNA world.[109]
Stop codons: Codons for translational stops are also an interesting aspect to the problem of the origin of the genetic code. As an example for addressing stop codon evolution, it has been suggested that the stop codons are such that they are most likely to terminate translation early in the case of aframe shift error.[110] In contrast, some stereochemical molecular models explain the origin of stop codons as "unassignable".[82]
^"The Nobel Prize in Physiology or Medicine 1959" (Press release). The Royal Swedish Academy of Science. 1959. Retrieved27 February 2010.The Nobel Prize in Physiology or Medicine 1959 was awarded jointly to Severo Ochoa and Arthur Kornberg 'for their discovery of the mechanisms in the biological synthesis of ribonucleic acid and deoxyribonucleic acid'.
^"The Nobel Prize in Physiology or Medicine 1968" (Press release). The Royal Swedish Academy of Science. 1968. Retrieved27 February 2010.The Nobel Prize in Physiology or Medicine 1968 was awarded jointly to Robert W. Holley, Har Gobind Khorana and Marshall W. Nirenberg 'for their interpretation of the genetic code and its function in protein synthesis'.
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