Abase pair (bp) is a fundamental unit of double-strandednucleic acids consisting of twonucleobases bound to each other byhydrogen bonds. They form the building blocks of theDNA double helix and contribute to the folded structure of both DNA andRNA. Dictated by specifichydrogen bonding patterns, "Watson–Crick" (or "Watson–Crick–Franklin") base pairs (guanine–cytosine andadenine–thymine)[1] allow the DNA helix to maintain a regular helical structure that is subtly dependent on itsnucleotide sequence.[2] Thecomplementary nature of this based-paired structure provides aredundant copy of thegenetic information encoded within each strand of DNA. The regular structure and data redundancy provided by the DNA double helix make DNA well suited to the storage of genetic information, while base-pairing between DNA and incoming nucleotides provides the mechanism through whichDNA polymerase replicates DNA andRNA polymerase transcribes DNA into RNA. Many DNA-binding proteins can recognize specific base-pairing patterns that identify particular regulatory regions of genes.
Intramolecular base pairs can occur within single-stranded nucleic acids. This is particularly important in RNA molecules (e.g.,transfer RNA), where Watson–Crick base pairs (guanine–cytosine and adenine–uracil) permit the formation of short double-stranded helices, and a wide variety of non–Watson–Crick interactions (e.g., G–U or A–A) allow RNAs to fold into a vast range of specific three-dimensionalstructures. In addition, base-pairing betweentransfer RNA (tRNA) andmessenger RNA (mRNA) forms the basis for themolecular recognition events that result in the nucleotide sequence of mRNA becomingtranslated into the amino acid sequence ofproteins via thegenetic code.
The size of an individualgene or an organism's entiregenome is often measured in base pairs because DNA is usually double-stranded. Hence, the number of total base pairs is equal to the number of nucleotides in one of the strands (with the exception of non-coding single-stranded regions oftelomeres). Thehaploidhuman genome (23chromosomes) is estimated to be about 3.2 billion base pairs long and to contain 20,000–25,000 distinct protein-coding genes.[3][4][5][6] Akilobase (kb) is a unit of measurement inmolecular biology equal to 1000 base pairs of DNA or RNA.[7] The total number ofDNA base pairs on Earth is estimated at 5.0×1037 with a weight of 50 billiontonnes.[8] In comparison, the totalmass of thebiosphere has been estimated to be as much as 4 TtC (trillion tons ofcarbon).[9]
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Hydrogen bonding is the chemical interaction that underlies the base-pairing rules described above. Appropriate geometrical correspondence of hydrogen bond donors and acceptors allows only the "right" pairs to form stably. DNA with highGC-content is more stable than DNA with low GC-content. Crucially, however,stacking interactions are primarily responsible for stabilising the double-helical structure; Watson-Crick base pairing's contribution to global structural stability is minimal, but its role in the specificity underlying complementarity is, by contrast, of maximal importance as this underlies the template-dependent processes of thecentral dogma (e.g.DNA replication).[10]
The biggernucleobases, adenine and guanine, are members of a class of double-ringed chemical structures calledpurines; the smaller nucleobases, cytosine and thymine (and uracil), are members of a class of single-ringed chemical structures calledpyrimidines. Purines are complementary only with pyrimidines: pyrimidine–pyrimidine pairings are energetically unfavorable because the molecules are too far apart for hydrogen bonding to be established; purine–purine pairings are energetically unfavorable because the molecules are too close, leading to overlap repulsion. Purine–pyrimidine base-pairing of AT or GC or UA (in RNA) results in proper duplex structure. The only other purine–pyrimidine pairings would be AC and GT and UG (in RNA); these pairings are mismatches because the patterns of hydrogen donors and acceptors do not correspond. The GU pairing, with two hydrogen bonds, does occur fairly often inRNA (seewobble base pair).
Paired DNA and RNA molecules are comparatively stable at room temperature, but the two nucleotide strands will separate above amelting point that is determined by the length of the molecules, the extent of mispairing (if any), and the GC content. Higher GC content results in higher melting temperatures; it is, therefore, unsurprising that the genomes ofextremophile organisms such asThermus thermophilus are particularly GC-rich. On the converse, regions of a genome that need to separate frequently — for example, the promoter regions for often-transcribed genes — are comparatively GC-poor (for example, seeTATA box). GC content and melting temperature must also be taken into account when designingprimers forPCR reactions.[citation needed]
The following DNA sequences illustrate pair double-stranded patterns. By convention, the top strand is written from the5′-end to the3′-end; thus, the bottom strand (complementary strand) is written 3′ to 5′.
ATCGATTGAGCTCTAGCGTAGCTAACTCGAGATCGCAUCGAUUGAGCUCUAGCGUAGCUAACUCGAGAUCGCChemical analogs of nucleotides can take the place of proper nucleotides and establish non-canonical base-pairing, leading to errors (mostlypoint mutations) inDNA replication andDNA transcription. This is due to theirisosteric chemistry. One common mutagenic base analog is5-bromouracil, which resembles thymine but can base-pair to guanine in itsenol form.[11]
Other chemicals, known asDNA intercalators, fit into the gap between adjacent bases on a single strand and induceframeshift mutations by "masquerading" as a base, causing the DNA replication machinery to skip or insert additional nucleotides at the intercalated site. Most intercalators are largepolyaromatic compounds and are known or suspectedcarcinogens. Examples includeethidium bromide andacridine.[12][citation needed]
Mismatched base pairs can be generated by errors ofDNA replication and as intermediates duringhomologous recombination. The process of mismatch repair ordinarily must recognize and correctly repair a small number of base mispairs within a long sequence of normal DNA base pairs. To repair mismatches formed during DNA replication, several distinctive repair processes have evolved to distinguish between the template strand and the newly formed strand so that only the newly inserted incorrect nucleotide is removed (in order to avoid generating a mutation).[13] The proteins employed in mismatch repair during DNA replication, and the clinical significance of defects in this process are described in the articleDNA mismatch repair. The process of mispair correction during recombination is described in the articlegene conversion.
The following abbreviations are commonly used to describe the length of a D/RNA molecule:
For single-stranded DNA/RNA, units ofnucleotides are used—abbreviated nt (or knt, Mnt, Gnt)—as they are not paired.To distinguish between units ofcomputer storage and bases, kbp, Mbp, Gbp, etc. may be used for base pairs.
Thecentimorgan is also often used to imply distance along a chromosome, but the number of base pairs it corresponds to varies widely. In the human genome, the centimorgan is about 1 million base pairs.[15][16]
An unnatural base pair (UBP) is a designed subunit (ornucleobase) ofDNA which is created in a laboratory and does not occur in nature. DNA sequences have been described which use newly created nucleobases to form a third base pair, in addition to the two base pairs found in nature, A-T (adenine –thymine) and G-C (guanine –cytosine). A few research groups have been searching for a third base pair for DNA, including teams led bySteven A. Benner,Philippe Marliere,Floyd E. Romesberg andIchiro Hirao.[17] Some new base pairs based on alternative hydrogen bonding, hydrophobic interactions and metal coordination have been reported.[18][19][20][21]
In 1989 Steven Benner (then working at theSwiss Federal Institute of Technology in Zurich) and his team led with modified forms of cytosine and guanine into DNA moleculesin vitro.[22] The nucleotides, which encoded RNA and proteins, were successfully replicatedin vitro. Since then, Benner's team has been trying to engineer cells that can make foreign bases from scratch, obviating the need for a feedstock.[23]
In 2002, Ichiro Hirao's group in Japan developed an unnatural base pair between 2-amino-8-(2-thienyl)purine (s) and pyridine-2-one (y) that functions in transcription and translation, for the site-specific incorporation of non-standard amino acids into proteins.[24] In 2006, they created 7-(2-thienyl)imidazo[4,5-b]pyridine (Ds) and pyrrole-2-carbaldehyde (Pa) as a third base pair for replication and transcription.[25] Afterward, Ds and 4-[3-(6-aminohexanamido)-1-propynyl]-2-nitropyrrole (Px) was discovered as a high fidelity pair in PCR amplification.[26][27] In 2013, they applied the Ds-Px pair to DNA aptamer generation byin vitro selection (SELEX) and demonstrated the genetic alphabet expansion significantly augment DNA aptamer affinities to target proteins.[28]
In 2012, a group of American scientists led by Floyd Romesberg, a chemical biologist at theScripps Research Institute in San Diego, California, published that his team designed an unnatural base pair (UBP).[20] The two new artificial nucleotides orUnnatural Base Pair (UBP) were namedd5SICS anddNaM. More technically, these artificialnucleotides bearing hydrophobicnucleobases, feature two fusedaromatic rings that form a (d5SICS–dNaM) complex or base pair in DNA.[23][29] His team designed a variety ofin vitro or "test tube" templates containing the unnatural base pair and they confirmed that it was efficiently replicated with high fidelity in virtually all sequence contexts using the modern standardin vitro techniques, namelyPCR amplification of DNA and PCR-based applications.[20] Their results show that for PCR and PCR-based applications, the d5SICS–dNaM unnatural base pair is functionally equivalent to a natural base pair, and when combined with the other two natural base pairs used by all organisms, A–T and G–C, they provide a fully functional and expanded six-letter "genetic alphabet".[29]
In 2014 the same team from the Scripps Research Institute reported that they synthesized a stretch of circular DNA known as aplasmid containing natural T-A and C-G base pairs along with the best-performing UBP Romesberg's laboratory had designed and inserted it into cells of the common bacteriumE. coli that successfully replicated the unnatural base pairs through multiple generations.[17] Thetransfection did not hamper the growth of theE. coli cells and showed no sign of losing its unnatural base pairs to its naturalDNA repair mechanisms. This is the first known example of a living organism passing along an expanded genetic code to subsequent generations.[29][30] Romesberg said he and his colleagues created 300 variants to refine the design of nucleotides that would be stable enough and would be replicated as easily as the natural ones when the cells divide. This was in part achieved by the addition of a supportivealgal gene that expresses anucleotide triphosphate transporter which efficiently imports the triphosphates of both d5SICSTP and dNaMTP intoE. coli bacteria.[29] Then, the natural bacterial replication pathways use them to accurately replicate aplasmid containing d5SICS–dNaM. Other researchers were surprised that the bacteria replicated these human-made DNA subunits.[31]
The successful incorporation of a third base pair is a significant breakthrough toward the goal of greatly expanding the number ofamino acids which can be encoded by DNA, from the existing 20 amino acids to a theoretically possible 172, thereby expanding the potential for living organisms to produce novelproteins.[17] The artificial strings of DNA do not encode for anything yet, but scientists speculate they could be designed to manufacture new proteins which could have industrial or pharmaceutical uses.[32] Experts said the synthetic DNA incorporating the unnatural base pair raises the possibility of life forms based on a different DNA code.[31][32]
In addition to the canonical pairing, some conditions can also favour base-pairing with alternative base orientation, and number and geometry of hydrogen bonds. These pairings are accompanied by alterations to the local backbone shape.[citation needed]
The most common of these is thewobble base pairing that occurs betweentRNAs andmRNAs at the third base position of manycodons duringtranscription[34] and during the charging of tRNAs by sometRNA synthetases.[35] They have also been observed in the secondary structures of some RNA sequences.[36]
Additionally,Hoogsteen base pairing (typically written as A•U/T and G•C) can exist in some DNA sequences (e.g. CA and TA dinucleotides) in dynamic equilibrium with standard Watson–Crick pairing.[33] They have also been observed in some protein–DNA complexes.[37]
In addition to these alternative base pairings, a wide range of base-base hydrogen bonding is observed in RNA secondary and tertiary structure.[38] These bonds are often necessary for the precise, complex shape of an RNA, as well as its binding to interaction partners.[38]
Each mutagenic event in the presence of an acridine results in the addition or removal of a single base pair.
...in humans 1 centimorgan on average represents a distance of about 7.5x105 base pairs.
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