<i>In Vitro</i>
            Selection of Functional Oligonucleotides and the Origins of Biochemical Activity

Carothers, James M.; Szostak, Jack W.

doi:10.1002/3527608192.ch1

Cited by 12 publications

(13 citation statements)

References 154 publications

(184 reference statements)

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“…This cycle is repeated several times (6 to 10) and as a result the DNA or RNA sequence of a high affinity to target ligand is obtained. The SELEX technique is discussed in detail in a number of papers and reviews (see, for example [8,10,11]). …”

Section: Introductionmentioning

confidence: 99%

Electrochemical Aptasensors – Recent Achievements and Perspectives

2009

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This article reviews recent achievements in developing aptamer-based electrochemical biosensors (electrochemical aptasensors). Aptamers are single stranded DNA or RNA molecules with high specificity to various ligands. Their specificity is comparable and in certain cases even higher than those of antibodies. In contrast to antibodies, aptamers are prepared by an in vitro selection procedure developed simultaneously in the early 1990s by L. Gold and A. Ellington. Due to their stability and the possibility of chemical modification aptamers can be immobilized on various supports and serve as artificial receptors in biosensors. The first aptasensors developed in the second half of 1990s were based on optical detection. However, in early 2000 substantial interest arose to the development of electrochemical aptasensors. It has been shown that due to their simplicity and fast response they represent an excellent tool in practical applications. The main focus of this review is to discuss the configuration of aptamers and electrochemical methods for detecting aptamers -analyte interactions. We will also provide a brief history of aptamer development, along with molecular structure and methods of aptamer engineering. Methods for immobilizing aptamers onto a solid support are also discussed.

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Section: Introductionmentioning

confidence: 99%

Electrochemical Aptasensors – Recent Achievements and Perspectives

2009

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“…Others have emphasized that a generalizable information-function relationship may be a natural consequence of the scale of monomer subunits in RNA relative to small molecule ligands or substrates (Lau et al 2004). With greater understanding of how and why information and function are related in RNA structures it may be possible to devise better methods for creating biosensors and therapeutics (for review, see Sullenger and Gilboa 2002;Rimmele 2003) and gain insight into the origins and evolution of biochemical activity (Carothers and Szostak 2006).…”

Section: Introductionmentioning

confidence: 99%

Solution structure of an informationally complex high-affinity RNA aptamer to GTP

Carothers¹,

Davis²,

Chou³

et al. 2006

RNA

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Higher-affinity RNA aptamers to GTP are more informationally complex than lower-affinity aptamers. Analog binding studies have shown that the additional information needed to improve affinity does not specify more interactions with the ligand. In light of those observations, we would like to understand the structural characteristics that enable complex aptamers to bind their ligands with higher affinity. Here we present the solution structure of the 41-nt Class I GTP aptamer (K d = 75 nM) as determined by NMR. The backbone of the aptamer forms a reverse-S that shapes the binding pocket. The ligand nucleobase stacks between purine platforms and makes hydrogen bonds with the edge of another base. Interestingly, the local modes of interaction for the Class I aptamer and an RNA aptamer that binds ATP with a K d of 6 mM are very much alike. The aptamers exhibit nearly identical levels of binding specificity and fraction of ligand sequestered from the solvent (81%-85%). However, the GTP aptamer is more informationally complex ($45 vs. 35 bits) and has a larger recognition bulge (15 vs. 12 nucleotides) with many more stabilizing base-base interactions. Because the aptamers have similar modes of ligand binding, we conclude that the stabilizing structural elements in the Class I aptamer are responsible for much of the difference in K d . These results are consistent with the hypothesis that increasing the number of intra-RNA interactions, rather than adding specific contacts to the ligand, is the simplest way to improve binding affinity.

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“…Synthetic RNA aptamers can be generated with in vitro selection to bind a chosen molecular target [8] and deployed as genetic control mechanisms [7]. In one successful example, RNA aptamers evolved to bind the tetracycline repressor protein (TetR) in E. coli induce gene expression from a tet promoter to the same extent as tetracycline [9].…”

Section: Dynamic Gene Regulation With Rnamentioning

confidence: 99%