Aptamer

Source: Wikipedia, the free encyclopedia.
(Redirected from
Aptamers
)
Left: Unbound aptamer. Right: the aptamer bound to its target protein. The protein is in yellow. Parts of the aptamer that change shape when it binds its target are in blue, while the unchanging parts are in orange. The parts of the aptamer that contact the protein are highlighted in red.
UV light. This type of test allows a doctor or researcher to identify cancer cells in a tissue sample from a patient
.

Aptamers are

proteins. This difference can make aptamers a better choice than antibodies for some purposes (see antibody replacement
).

Aptamers are used in biological lab research and

drug delivery systems and controlled drug release systems. They also find use in other molecular engineering
tasks.

Most aptamers originate from

mutate or change the chemistry of the aptamers and do another selection, or might use rational design processes to engineer improvements. Non-SELEX methods
for discovering aptamers also exist.

Researchers optimize aptamers to achieve a variety of beneficial features. The most important feature is

speed of binding. As the yield of the synthesis used to produce known aptamers shrinks quickly for longer sequences,[4]
researchers often truncate aptamers to the minimal binding sequence to reduce the production cost.

Etymology

The word "aptamer" is a neologism coined by Andrew Ellington and Jack Szostak in their first publication on the topic. They did not provide a precise definition, stating "We have termed these individual RNA sequences 'aptamers', from the Latin 'aptus', to fit."[5]

The word itself, however, derives from the Greek word ἅπτω, to connect or fit (as used by Homer (c. 8th century BC)[6] [7]) and μέρος, a component of something larger.[8]

Classification

A typical aptamer is a synthetically generated ligand exploiting the combinatorial diversity of DNA, RNA, XNA, or peptide to achieve strong, specific binding for a particular target molecule or family of target molecules. Aptamers are occasionally classified as "chemical antibodies" or "antibody mimics".[9] However, most aptamers are small, with a molecular weight of 6-30 kDa, in contrast to the 150 kDa size of antibodies, and contain one binding site rather than the two matching antigen binding regions of a typical antibody.

History

Jack Szostak, Nobel laureate and one of the inventors of SELEX and aptamers.

Since its first application in 1967,[10] directed evolution methodologies have been used to develop biomolecules with new properties and functions. Early examples include the modification of the bacteriophage Qbeta replication system and the generation of ribozymes with modified cleavage activity.[11]

In 1990, two teams independently developed and published SELEX (Systematic Evolution of Ligands by EXponential enrichment) methods and generated RNA aptamers: the lab of Larry Gold, using the term SELEX for their process of selecting RNA

organic dyes.[5][13] Two years later, the Szostak lab and Gilead Sciences, acting independently of one another, used in vitro selection schemes to generate DNA aptamers for organic dyes[14] and human thrombin,[15] respectively. In 2001, SELEX was automated by J. Colin Cox in the Ellington lab, reducing the duration of a weeks-long selection experiment to just three days.[16][17][18]

In 2002, two groups led by

origin of life on Earth.[19]

Properties

Structure

The complex and diverse secondary and tertiary structure of aptamers, as shown in this schematic of an aptamer's secondary structure, is what lets them bind their target strongly and specifically. Complementary base pairing is visible in the black lines connecting some G-C and A-T bases. This is a feature of nucleic acids that does not exist in the amino acids of antibodies. It helps aptamers form these unique structures. Hairpin regions (red), which rely on this base pairing, enhance the aptamer's stability at different temperatures. This image also shows examples of chemical modifications to the base aptamer. Two unnatural bases, which enhance durability, are in yellow. The biotin molecule binds with extreme strength to streptavidin, allowing the aptamer to be anchored to other molecules or to a surface in sensors and assays.

Most aptamers are based on a specific oligomer sequence of 20-100 bases and 3-20 kDa. Some have chemical modifications for functional enhancements or compatibility with larger engineered molecular systems. DNA, RNA, XNA, and peptide aptamer chemistries can each offer distinct profiles in terms of shelf stability, durability in serum or in vivo, specificity and sensitivity, cost, ease of generation, amplification, and characterization, and familiarity to users. Typically, DNA- and RNA-based aptamers exhibit low immunogenicity, are amplifiable via Polymerase Chain Reaction (PCR), and have complex secondary structure and tertiary structure.[20][21][22][23] DNA- and XNA-based aptamers exhibit superior shelf stability. XNA-based aptamers can introduce additional chemical diversity to increase binding affinity or greater durability in serum or in vivo.

As 22 genetically-encoded and over 500 naturally-occurring

amino acids exist, peptide aptamers, as well as antibodies, have much greater potential combinatorial diversity per unit length relative to the 4 nucleic acids in DNA or RNA.[24] Chemical modifications of nucleic acid bases or backbones increase the chemical diversity of standard nucleic acid bases.[25]

Split aptamers are composed of two or more DNA strands that are pieces of a larger parent aptamer that has been broken in two by a molecular nick.[26] The ability of each component strand to bind targets will depend on the location of the nick, as well as the secondary structures of the daughter strands.[27] The presence of a target molecule supports the DNA fragments joining together. This can be used as the basis for biosensors.[28] Once assembled, the two separate DNA strands can be ligated into a single strand.

Unmodified aptamers are cleared rapidly from the

diagnostic imaging.[30]

In a study on aptamers[31] designed to bind with proteins associated with Ebola infection, a comparison was made among three aptamers isolated for their ability to bind the target protein EBOV sGP. Although these aptamers vary in both sequence and structure, they exhibit remarkably similar relative affinities for sGP from EBOV and SUDV, as well as EBOV GP1.2. Notably, these aptamers demonstrated a high degree of specificity for the GP gene products. One aptamer, in particular, proved effective as a recognition element in an electrochemical sensor, enabling the detection of sGP and GP1.2 in solution, as well as GP1.2 within a membrane context.The results of this research point to the intriguing possibility that certain regions on protein surfaces may possess aptatropic qualities. Identifying the key features of such sites, in conjunction with improved 3-D structural predictions for aptamers, holds the potential to enhance the accuracy of predicting aptamer interaction sites on proteins. This, in turn, may help identify aptamers with a heightened likelihood of binding proteins with high affinity, as well as shed light on protein mutations that could significantly impact aptamer binding.This comprehensive understanding of the structure-based interactions between aptamers and proteins is vital for refining the computational predictability of aptamer-protein binding. Moreover, it has the potential to eventually eliminate the need for the experimental SELEX protocol.

Targets

Aptamer targets can include small molecules and

prostate specific antigen (PSA),[42][43] dopamine,[44] and the non-classical oncogene, heat shock factor 1 (HSF1).[45]

Aptamers have been generated against cancer cells,

SARS coronavirus (SARS-CoV)[49] and SARS-CoV-2.[50]

Aptamers may be particularly useful for

a-amanitin (the toxin that causes lethal Amanita poisoning) has been developed, an example of an aptamer against a mushroom target.[55]

Aptamer applications can be roughly grouped into sensing, therapeutic, reagent production, and engineering categories. Sensing applications are important in environmental, biomedical,

precision medicine, aptamers can function as drugs,[61] as targeted drug delivery vehicles,[62] as controlled release mechanisms, and as reagents for drug discovery via high-throughput screening for small molecules[63] and proteins.[64][65] Aptamers have application for protein production monitoring, quality control, and purification.[66][67][68] They can function in molecular engineering applications as a way to modify proteins, such as enhancing DNA polymerase to make PCR more reliable.[69][70][71][72]

Because the affinity of the aptamer also affects its dynamic range and limit of detection, aptamers with a lower affinity may be desirable when assaying high concentrations of a target molecule.[73] Affinity chromatography also depends on the ability of the affinity reagent, such as an aptamer, to bind and release its target, and lower affinities may aid in the release of the target molecule.[74] Hence, specific applications determine the useful range for aptamer affinity.

Antibody replacement

Aptamers can replace antibodies in many

replicability and lower cost, a greater level of control due to the in vitro selection conditions, and capacity to be efficiently engineered for durability, specificity, and sensitivity.[82]

In addition, aptamers contribute to reduction of research animal use.[83] While antibodies often rely on animals for initial discovery, as well as for production in the case of polyclonal antibodies, both the selection and production of aptamers is typically animal-free. However, phage display methods allow for selection of antibodies in vitro, followed by production from a monoclonal cell line, avoiding the use of animals entirely.[84]

Controlled release of therapeutics

The ability of aptamers to reversibly bind molecules such as proteins has generated increasing interest in using them to facilitate

controlled release of therapeutic biomolecules, such as growth factors. This can be accomplished by tuning the binding strength to passively release the growth factors,[85] along with active release via mechanisms such as hybridization of the aptamer with complementary oligonucleotides[86] or unfolding of the aptamer due to cellular traction forces.[87]

AptaBiD

AptaBiD (Aptamer-Facilitated Biomarker Discovery) is an aptamer-based method for biomarker discovery.[88]

Peptide Aptamers

While most aptamers are based on DNA, RNA, or XNA, peptide aptamers[89] are artificial proteins selected or engineered to bind specific target molecules.

Structure

Peptide aptamers consist of one or more peptide loops of variable sequence displayed by a protein scaffold. Derivatives known as tadpoles, in which peptide aptamer "heads" are covalently linked to unique sequence double-stranded DNA "tails", allow quantification of scarce target molecules in mixtures by PCR (using, for example, the quantitative real-time polymerase chain reaction) of their DNA tails.[90] The peptides that form the aptamer variable regions are synthesized as part of the same polypeptide chain as the scaffold and are constrained at their N and C termini by linkage to it. This double structural constraint decreases the diversity of the 3D structures that the variable regions can adopt,[91] and this reduction in structural diversity lowers the entropic cost of molecular binding when interaction with the target causes the variable regions to adopt a uniform structure.

Selection

The most common peptide aptamer selection system is the

two-hybrid system. Peptide aptamers can also be selected from combinatorial peptide libraries constructed by phage display and other surface display technologies such as mRNA display, ribosome display, bacterial display and yeast display. These experimental procedures are also known as biopanning. All the peptides panned from combinatorial peptide libraries have been stored in the MimoDB database.[92][93]

Applications

Libraries of peptide aptamers have been used as "mutagens", in studies in which an investigator introduces a library that expresses different peptide aptamers into a cell population, selects for a desired phenotype, and identifies those aptamers that cause the phenotype. The investigator then uses those aptamers as baits, for example in yeast two-hybrid screens to identify the cellular proteins targeted by those aptamers. Such experiments identify particular proteins bound by the aptamers, and protein interactions that the aptamers disrupt, to cause the phenotype.[94][95] In addition, peptide aptamers derivatized with appropriate functional moieties can cause specific post-translational modification of their target proteins, or change the subcellular localization of the targets.[96]

This assay tests the ability of two different types of aptamers (V and I) to detect their respective protein targets (VEGF and IFN-y). The labels Apt1, Apt2, Apt3, and Apt4 are in decreasing order of binding strength (i.e. Apt1 is the strongest aptamer). The DD, AD, DA, and AA letters mean that they have different combinations of unnatural base pairs. This causes their difference in binding strengths. The "-" columns have no protein, and the "+" columns do have protein. Aptamer with protein (+) and without protein (-) is loaded into wells in a gel and moves down the column lanes. If target is present, they bind and travel more slowly, due to the charge on the aptamer and the mass of the protein. Otherwise, the unbound aptamer moves quickly to the end of the lane. The difference in position between the "+" and "-" bands is the "mobility shift." This allows the researcher to detect the presence or absence of the protein. The darker band in the leftmost V and I lanes show that stronger aptamer-target binding makes it easier to detect the target at a given amount of target protein in the sample. The bottom image includes denaturing urea in the gel that disrupts aptamer-target binding in the weaker I aptamers, showing that the aptamer-protein binding is indeed what caused the mobility shift.

Industry and Research Community

Commercial products and companies based on aptamers include the drug Macugen (pegaptanib)[97] and the clinical diagnostic company SomaLogic.[98] The International Society on Aptamers (INSOAP), a professional society for the aptamer research community, publishes a journal devoted to the topic, Aptamers. Apta-index[99] is a current database cataloging and simplifying the ordering process for over 700 aptamers.

See also

References

  1. S2CID 36797240
    .
  2. PMID 23326761
    .
  3. PMID 27155114
    .
  4. ^ "DNA Oligonucleotide Synthesis". Millipore Sigma. Retrieved 4 July 2022.
  5. ^
    S2CID 4273647
    .
  6. ^ "ἅπτω", Βικιλεξικό (in Greek), 2023-03-12, retrieved 2024-03-21
  7. ^ "Οδύσσεια/φ - Βικιθήκη". el.wikisource.org (in Greek). Retrieved 2024-03-21.
  8. ^ "μέρος", Wiktionary, the free dictionary, 2023-05-31, retrieved 2024-03-21
  9. S2CID 16618423
    .
  10. PMID 5231602
    .
  11. PMID 2684778
    .
  12. PMID 2200121
    .
  13. PMID 17627883
    .
  14. S2CID 4332485
    .
  15. S2CID 4349607
    .
  16. PMID 11557339
    .
  17. ^ Cox JC, Rajendran M, Riedel T, Davidson EA, Sooter LJ, Bayer TS, et al. (June 2002). "Automated acquisition of aptamer sequences". Combinatorial Chemistry & High Throughput Screening. 5 (4): 289–299.
    PMID 12052180
    .
  18. ^ Cox JC, Hayhurst A, Hesselberth J, Bayer TS, Georgiou G, Ellington AD (October 2002). "Automated selection of aptamers against protein targets translated in vitro: from gene to aptamer". Nucleic Acids Research. 30 (20): 108e–108.
    PMID 12384610
    .
  19. PMID 21106649
    .
  20. S2CID 240281828
    .
  21. PMID 21035241
    .
  22. PMID 10926496
    .
  23. S2CID 221833821
    .
  24. PMID 22213382
    .
  25. PMID 27715478
    .
  26. doi:10.1016/j.trac.2016.04.006
    .
  27. PMID 24033257
    .
  28. PMID 32112111
    .
  29. PMID 26757406
    .
  30. PMID 24525205
    .
  31. ^ Banerjee, S.; Hemmat, M.A.; Shubham, S.; Gosai, A.; Devarakonda, S.; Jiang, N.; Geekiyanage, C.; Dillard, J.A.; Maury, W.; Shrotriya, P.; et al. Structurally Different Yet Functionally Similar: Aptamers Specific for the Ebola Virus Soluble Glycoprotein and GP1,2 and Their Application in Electrochemical Sensing. Int. J. Mol. Sci. 2023, 24, 4627. https://doi.org/10.3390/ijms24054627
  32. ^
    PMID 36131986
    .
  33. PMID 28146093
    .
  34. S2CID 23670
    .
  35. PMID 18971322
    .
  36. PMID 37531184
    .
  37. PMID 17002307
    .
  38. doi:10.1246/bcsj.82.99
    .
  39. PMID 18406597
    .
  40. S2CID 8833436
    .
  41. S2CID 80613434
    .
  42. PMID 20692149
    .
  43. S2CID 22201181
    .
  44. PMID 19699181
    .
  45. PMID 24800749
    .
  46. S2CID 37058942
    .
  47. S2CID 36801266
    .
  48. PMID 28728009
    .
  49. ^
    PMID 30577479
    .
  50. PMID 33683787
    .
  51. PMID 32260091
    .
  52. ^
    PMID 31801185
    .
  53. PMID 29351868
    .
  54. S2CID 20781208
    .
  55. S2CID 3638299
    .
  56. ^ Penner G (July 2012). "Commercialization of an aptamer-based diagnostic test" (PDF). NeoVentures.
  57. PMID 17851611
    .
  58. PMID 26866998
    .
  59. PMID 21860458
    .
  60. doi:10.1016/j.bios.2018.10.010
    .
  61. PMID 33050158
    .
  62. S2CID 53562925
    .
  63. S2CID 4997899
    .
  64. S2CID 234061584
    .
  65. PMID 32796729
    .
  66. doi:10.1016/j.trac.2017.07.023
    . Retrieved 4 July 2022.
  67. PMID 12954786
    .
  68. S2CID 219564070
    .
  69. S2CID 244954278
    .
  70. S2CID 13406870
    .
  71. PMID 22865679
    .
  72. S2CID 243998479
    .
  73. PMID 32402748
    .
  74. ISBN 978-1-118-00219-3
    .
  75. PMID 25912679
    .
  76. PMID 25278480
    .
  77. ^ Bruno JG, Sivils JC (2016). "Aptamer "Western" blotting for E. coli outer membrane proteins and key virulence factors in pathogenic E. coli serotypes". Aptamers and Synthetic Antibodies.
  78. PMID 26862683
    .
  79. S2CID 13996688
    .
  80. PMID 27807347
    .
  81. PMID 25913927
    .
  82. S2CID 53242220
    .
  83. S2CID 1002312
    .
  84. PMID 32983137
    .
  85. PMID 20198232
    .
  86. PMID 22816442
    .
  87. PMID 30614086
    .
  88. PMID 18558676
    .
  89. S2CID 4327303
    .
  90. S2CID 1423778
    .
  91. PMID 8303294
    .
  92. PMID 22053087
    .
  93. ^ "MimoDB: a mimotope database and beyond". immunet.cn. Archived from the original on 2012-11-16. Retrieved 2016-02-03.
  94. PMID 10411916
    .
  95. PMID 23579184
    .
  96. PMID 11106396
    .
  97. ^ "FDA Approves Eyetech/Pfizer's Macugen". Review of Ophthalmology. Retrieved 30 June 2022.
  98. ^ Dutt S. "SomaLogic and Illumina Combine Strengths to Propel Innovation in Proteomics". BioSpace. Retrieved 30 June 2022.
  99. ^ "Apta-Index™ (Aptamer Database) - Library of 500+ Aptamers". APTAGEN, LLC. Retrieved 2022-12-16.

Further reading

Retrieved from "https://en.wikipedia.org/w/index.php?title=Aptamer&oldid=1215464127"