Rapid method for detection of point mutations using mismatch binding protein (MutS) and an optical biosensor

Gotoh, Masao; Hasebe, Masaharu; Ohira, Tomoko; Hasegawa, Yukio; Shinohara, Yasuro; Sota, Hiroyuki; Nakao, Junko; Tosu, Mariko

doi:10.1016/s1050-3862(97)00009-0

Cited by 45 publications

(46 citation statements)

References 11 publications

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“…The use of a biotin-SA linker for immobilization of oligonucleotides for studies of protein:DNA binding using columns and gels has been well proven in a number of experiments to allow the immobilized dsDNAs to remain active and selective in their protein binding [7,[12][13][14][15][16][17][18][19][20][21][22][23]. To our knowledge this is the first demonstration d the use of a biotin-SA linker for immobilization of oligonucleotides to study protein:DNA binding with a planar dsDNA monolayer.…”

Section: Discussionmentioning

confidence: 99%

<title>Probing protein: DNA interactions using a uniform monolayer of DNA and surface plasmon resonance</title>

Shumaker‐Parry¹,

Campbell²,

Stormo³

et al. 2000

SPIE Proceedings

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A method is described for immobilizing double-stranded DNAs to a planar gold surface with high density (1-3x10'2 DNA/cm2, depending on their length) and uniform spacing (-4 nm closest possible DNA-DNA separation). This is accomplished by adsorbing biotinylated DNAs onto a nearly close-packed monolayer of the protein streptavidin. This streptavidin monolayer, which offers -5x1O'2 biotin sites per cm2, is prepared first by adsorbing it onto a mixed sell-assembled monolayer on gold which contains biotin-terminated and oligo(ethylene glycol)-terminated alkylthiolates in a 3/7 ratio. This DNA-functionalized surface resists non-specific protein adsorption and is useful for probing the kinetics and equilibrium binding of proteins to DNA with surface plasmon resonance. This is demonstrated with the Mnt protein, which is found to bind in 3.8:1 ratio to its immobilized DNA operator sequence. This is consistent with its behavior in homogeneous solution. where it binds as a tetramer to its DNA. A sequence with a single base-pair mutation shows nearly as much Mnt binding, but a completely random DNA sequence shows only 5% of this binding. This proves that DNA-binding proteins bind sequence-specifically to double-stranded DNAs which are immobilized to gold with this streptavidin linker layer.

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Section: Discussionmentioning

confidence: 99%

<title>Probing protein: DNA interactions using a uniform monolayer of DNA and surface plasmon resonance</title>

Shumaker‐Parry¹,

Campbell²,

Stormo³

et al. 2000

SPIE Proceedings

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“…[3][4][5][6] SPR is attractive owing to several inherent advantages (e.g., labelfree analysis, simplicity, cost-effectiveness, and high sensitivity). [7][8][9][10][11][12][13][14][15][16][17][18][19][20][21][22][23][24] The SPR resonance angle variation is dependent on various physicochemical processes occurring at the SPR substrate (a thin metal film or a chemically modified metal surface) and/or the substrate/ solution interface. Thus, when coupled with electrochemistry (EC-SPR), it offers a viable avenue to probe optical and electrochemical properties of adsorbates and a sensitive means to quantify thickness variations of ultrathin films accompanying redox reactions.…”

Section: Introductionmentioning

confidence: 99%

Electrochemical Surface Plasmon Resonance Spectroscopy at Bilayered Silver/Gold Films

et al. 2006

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Bilayered silver/gold films (gold deposited on top of the silver film) were used as substrates for electrochemical surface plasmon resonance spectroscopy (EC-SPR). EC-SPR responses of electrochemical deposition/stripping of copper and redox-induced conformation changes of cytochrome c immobilized onto self-assembled monolayers preformed at these substrates were measured. Influence of the Ag layer thickness and the double-layer capacitance on the EC-SPR behavior was investigated. The results demonstrated that the bilayered Ag/Au metal films produce a sharper SPR dip profile than pure Au films and retain the high chemical stability of Au films. Contrary to the result by the Fresnel calculation that predicts a greater fraction of Ag in the bilayered film should result in a greater signal-to-noise ratio, the EC-SPR sensitivity is dependent on both the Ag/Au thickness ratio and the chemical modification of the surface. Factors affecting the overall SPR sensitivity at the bilayered films, such as the film morphology, potential-induced excess surface charges, and the adsorbate layer were investigated. Forming a compact adsorbate layer at the bilayered film diminishes the effect of potential-induce excess surface charges on the SPR signal and improves the overall EC-SPR sensitivity. For the case of redox-induced conformation changes of cytochrome c, the SPR signal obtained at the bilayered silver/gold film is 2.7 times as high as that at a pure gold film.

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“…The inability to detect mismatches using MutS protein in these experiments indicates that when MutS was used alone it was binding either to the ends of duplexes formed by hybridisation or to single stranded free probe, as was observed in previous studies (Gotoh et al, 1997, Babic et al, 1996. The second explanation would seem the more likely since the average signal obtained with MutS applied to complementary oligonucleotide was higher than when a mismatched sequence was used.…”

Section: Discussionmentioning

confidence: 59%

A novel optical biosensor format for the detection of clinically relevant TP53 mutations☆

Wilson¹,

Jiang²,

Minunni³

et al. 2005

Biosensors and Bioelectronics

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The TP53 gene has been the subject of intense research since the realisation that inactivation of this gene is common to most cancer types. Numerous publications have linked TP53 mutations in general or at specific locations to patient prognosis and therapy response. The findings of many studies using general approaches such as immunohistochemistry or sequencing are contradictory. However, the detection of specific mutations, especially those occurring in the structurally important L2 and L3 zinc binding domains, which are the most common sites of TP53 mutations, have been linked to patient prognosis and more strongly to radiotherapy and chemotherapy resistance in several major cancers. In this study, the TI-SPR-1 surface plasmon resonance system and Texas Instruments Spreeta chips were used to develop a DNA biosensor based on thiolated probes complementary to these domains. The sensors were able to detect these mutations in both oligonucleotides and PCR products with normal and mutant TP53 DNA, but the difference in hybridisation signal was small. Preliminary experiments to enhance the signal using Escherichia coli mismatch repair proteins, MutS and single strand binding protein were carried out. It was found that MutS was unable to bind to mismatch oligonucleotides, but single strand binding protein was able to bind to single stranded probes, which had not hybridised to the target, resulting in a three-fold increase in the sensitivity of the biosensor. While further work needs to be carried out to optimise the system, these preliminary experiments indicate that the TI-SPR-1 can be used for the detection of clinically relevant mutations in the TP53 gene and that the sensitivity can be increased significantly using single strand binding protein. This system has a number of advantages over current mutation detection technologies, including lower cost, ease of sensor preparation and measurement procedures, technical simplicity and increased speed due to the lack of need for gel electrophoresis.

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Rapid method for detection of point mutations using mismatch binding protein (MutS) and an optical biosensor

Cited by 45 publications

References 11 publications

<title>Probing protein: DNA interactions using a uniform monolayer of DNA and surface plasmon resonance</title>

<title>Probing protein: DNA interactions using a uniform monolayer of DNA and surface plasmon resonance</title>

Electrochemical Surface Plasmon Resonance Spectroscopy at Bilayered Silver/Gold Films

A novel optical biosensor format for the detection of clinically relevant TP53 mutations☆

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