In vitro selection and evaluation of rna aptamers that recognize arginine-rich-motif model peptide on a quartz-crystal microbalance

Fukusho, Shinobu; Furusawa, Hiroyuki; Okahata, Yoshio

doi:10.1039/b108940b

Cited by 21 publications

(16 citation statements)

References 10 publications

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“…We quantify the distribution of associated secondary-structural motifs using tree graphs and graph theory measures of size and complexity (particularly, tree diameter and second smallest Laplacian eigenvalue, which are defined in the Materials and Methods section). Our results show that such random RNA pools heavily favor (>90%) simple topological structures like linear stem-loop and low-branching motifs, as confirmed by many known aptamers (Bae et al 2002;Fukusho et al 2002;Khoo et al 2002;Komatsu et al 2002;Ulrich et al 2002;Zinnen et al 2002;Meli et al 2003;Laserson et al 2004). For example, the 25-nt and 40-nt pools are almost completely populated by hairpin and stem-bulge-stem-loop structures.…”

Section: Introductionmentioning

confidence: 61%

“…Indeed, in vitro selection experiments have revealed novel functional RNAs from random sequence pools, including a wide range of RNA aptamers that bind to particular compounds, such as ATP, antibiotics or proteins, and catalytic RNAs (Wilson and Szostak 1999;Hodgson and Suga 2004); see the collection of several hundred aptamers in the Aptamer Database (http:// aptamer.icmb.utexas.edu). However, an analysis of existing aptamers has shown that RNA sequences isolated from selection experiments tend to have simple topologies (Bae et al 2002;Fukusho et al 2002;Khoo et al 2002;Komatsu et al 2002;Ulrich et al 2002;Zinnen et al 2002;Meli et al 2003;Laserson et al 2004). That is, linear or slightly branched structures occur far more often than highly compact structures.…”

Section: Introductionmentioning

confidence: 99%

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In vitro RNA random pools are not structurally diverse: A computational analysis

Gevertz¹,

Gan²,

Schlick³

2005

RNA

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In vitro selection of functional RNAs from large random sequence pools has led to the identification of many ligand-binding and catalytic RNAs. However, the structural diversity in random pools is not well understood. Such an understanding is a prerequisite for designing sequence pools to increase the probability of finding complex functional RNA by in vitro selection techniques. Toward this goal, we have generated by computer five random pools of RNA sequences of length up to 100 nt to mimic experiments and characterized the distribution of associated secondary structural motifs using sets of possible RNA tree structures derived from graph theory techniques. Our results show that such random pools heavily favor simple topological structures: For example, linear stem-loop and low-branching motifs are favored rather than complex structures with high-order junctions, as confirmed by known aptamers. Moreover, we quantify the rise of structural complexity with sequence length and report the dominant class of tree motifs (characterized by vertex number) for each pool. These analyses show not only that random pools do not lead to a uniform distribution of possible RNA secondary topologies; they point to avenues for designing pools with specific simple and complex structures in equal abundance in the goal of broadening the range of functional RNAs discovered by in vitro selection. Specifically, the optimal RNA sequence pool length to identify a structure with x stems is 20x.

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Section: Introductionmentioning

confidence: 61%

Section: Introductionmentioning

confidence: 99%

In vitro RNA random pools are not structurally diverse: A computational analysis

Gevertz¹,

Gan²,

Schlick³

2005

RNA

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“…[39,40] To prevent any nonspecific interactions on the electrode surface during measurement, PEG molecules known to inhibit nonspecific interactions were used as a spacer to immobilize the KIX peptide. [41] The degree of immobilization at each step was confirmed directly from frequency decreases (mass increases) by using the QCM in water.…”

Section: Preparation Of the Kid Peptidementioning

confidence: 99%

Kinetic Study of Phosphorylation‐Dependent Complex Formation between the Kinase‐Inducible Domain (KID) of CREB and the KIX Domain of CBP on a Quartz Crystal Microbalance

Matsuno

Furusawa

Okahata

2004

Chemistry A European J

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We report quantitative analysis of peptide-peptide interactions on a 27 MHz quartz crystal microbalance (QCM) in aqueous solution. The KID (kinase-inducible domain) of transcription factor CREB (cyclic AMP response element binding protein) is known to interact with the KIX domain of coactivator CBP (CREB binding protein), facilitated by phosphorylation at Ser-133 of the KID. The KIX domain peptide (86 aa) was immobilized on the QCM gold electrode surface by means of a poly(ethylene glycol) spacer. Binding of the KID peptide (46 aa) to the KIX peptide was detected by frequency decreases (mass increases) of the QCM. Both maximum binding amount (Deltammax) and association constants (Ka) obtained from the QCM measurements increased as a result of phosphorylation of Ser-133 of the KID peptide. The Ka values for KIX peptide to the phosphorylated (pKID) and unphosphorylated KID peptides were (93+/-2) x 10(3) and (5+/-1) x 10(3) M(-1), respectively. This difference was explained by the dissociation rate constant (k(-1)) of the pKID being 20 times smaller than that of the KID, while association rate constants (k1) were independent of phosphorylation.

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“…Matsuno et al (2001a) designed and synthesized specific helical peptides on the basis of a well-conserved sequence of natural DNA-binding proteins and used these to investigate the sequence specificity and recognition rules of the DNA-binding proteins. A novel variation on the DNA biosensor was presented by Fukusho et al (2002) who used a peptide immobilized QCM to study RNA-peptide interactions. They conclude that five arginines were not enough to bind specifically to particular internal loops, but played an important role in recognizing specific RNAs.…”

Section: Dna-protein Interactionsmentioning

confidence: 99%

A survey of the 2001 to 2005 quartz crystal microbalance biosensor literature: applications of acoustic physics to the analysis of biomolecular interactions

Cooper¹,

Singleton²

2007

J of Molecular Recognition

335

205

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The widespread exploitation of biosensors in the analysis of molecular recognition has its origins in the mid-1990s following the release of commercial systems based on surface plasmon resonance (SPR). More recently, platforms based on piezoelectric acoustic sensors (principally 'bulk acoustic wave' (BAW), 'thickness shear mode' (TSM) sensors or 'quartz crystal microbalances' (QCM)), have been released that are driving the publication of a large number of papers analysing binding specificities, affinities, kinetics and conformational changes associated with a molecular recognition event. This article highlights salient theoretical and practical aspects of the technologies that underpin acoustic analysis, then reviews exemplary papers in key application areas involving small molecular weight ligands, carbohydrates, proteins, nucleic acids, viruses, bacteria, cells and lipidic and polymeric interfaces. Key differentiators between optical and acoustic sensing modalities are also reviewed.

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In vitro selection and evaluation of rna aptamers that recognize arginine-rich-motif model peptide on a quartz-crystal microbalance

Abstract: To study RNA-peptide interactions, we performed an in vitro selection of RNA on a simple alpha-helical peptide-immobilized quartz-crystal microbalance (QCM) and evaluated the association constants (10(7) M-1) of the selected RNA to the model peptide on the same QCM plate.

Cited by 21 publications

References 10 publications

In vitro RNA random pools are not structurally diverse: A computational analysis

In vitro RNA random pools are not structurally diverse: A computational analysis

Kinetic Study of Phosphorylation‐Dependent Complex Formation between the Kinase‐Inducible Domain (KID) of CREB and the KIX Domain of CBP on a Quartz Crystal Microbalance

A survey of the 2001 to 2005 quartz crystal microbalance biosensor literature: applications of acoustic physics to the analysis of biomolecular interactions

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