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Comprehensive characterization of molecular interactions based on nanomechanics.

Ghatkesar MK, Lang HP, Gerber C, Hegner M, Braun T - PLoS ONE (2008)

Bottom Line: Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro.This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology.The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: National Center of Competence for Research in Nanoscience, Institute of Physics, University of Basel, Basel, Switzerland.

ABSTRACT
Molecular interaction is a key concept in our understanding of the biological mechanisms of life. Two physical properties change when one molecular partner binds to another. Firstly, the masses combine and secondly, the structure of at least one binding partner is altered, mechanically transducing the binding into subsequent biological reactions. Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro. This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology. To prove the concept we measured lipid vesicle (approximately 748*10(6) Da) adsorption on the sensor interface followed by subsequent binding of the bee venom peptide melittin (2840 Da) to the vesicles. The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

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Related in: MedlinePlus

Combined mode measurements of vesicle and melittin adsorption on the cantilever sensor.The positive controls were pre-functionalized in such a way that vesicles only bind to the upper cantilever surface and melittin does not bind at all. The lower graph displays the mass adsorption and the upper graph reveals the surface stress development measured simultaneously. Note that the surface stress represents the differential signal between the positively and negatively functionalized cantilevers (two cantilevers each). The experiment was performed in 11 sections: (I) Baseline recording in buffer. (II, VI, VIII) Injections of 500 ng/ml DOPC vesicles. (III, V, VII, IX, XI) Buffer injections. (IV, X) Melittin injections (1 µM). Note that during the injection of melittin, the adsorbed mass is initially underestimated due to the high friction of the protein-solution, see also Braun et al., 2005 [5] for a discussion. During the subsequent buffer injection the correct mass is measured.
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pone-0003610-g002: Combined mode measurements of vesicle and melittin adsorption on the cantilever sensor.The positive controls were pre-functionalized in such a way that vesicles only bind to the upper cantilever surface and melittin does not bind at all. The lower graph displays the mass adsorption and the upper graph reveals the surface stress development measured simultaneously. Note that the surface stress represents the differential signal between the positively and negatively functionalized cantilevers (two cantilevers each). The experiment was performed in 11 sections: (I) Baseline recording in buffer. (II, VI, VIII) Injections of 500 ng/ml DOPC vesicles. (III, V, VII, IX, XI) Buffer injections. (IV, X) Melittin injections (1 µM). Note that during the injection of melittin, the adsorbed mass is initially underestimated due to the high friction of the protein-solution, see also Braun et al., 2005 [5] for a discussion. During the subsequent buffer injection the correct mass is measured.

Mentions: Supplemental Figure S2 (panel A) depicts the workflow of the main experiments and the results are shown in Figure 2. The adsorption of lipid to the cantilever has to be controlled carefully since asymmetrical functionalization of the cantilevers was crucial for detection of the static cantilever bending (a single sided coating was not a prerequisite for the mass adsorption signal). This was achieved by a specific pre-functionalization of the sensor interfaces.


Comprehensive characterization of molecular interactions based on nanomechanics.

Ghatkesar MK, Lang HP, Gerber C, Hegner M, Braun T - PLoS ONE (2008)

Combined mode measurements of vesicle and melittin adsorption on the cantilever sensor.The positive controls were pre-functionalized in such a way that vesicles only bind to the upper cantilever surface and melittin does not bind at all. The lower graph displays the mass adsorption and the upper graph reveals the surface stress development measured simultaneously. Note that the surface stress represents the differential signal between the positively and negatively functionalized cantilevers (two cantilevers each). The experiment was performed in 11 sections: (I) Baseline recording in buffer. (II, VI, VIII) Injections of 500 ng/ml DOPC vesicles. (III, V, VII, IX, XI) Buffer injections. (IV, X) Melittin injections (1 µM). Note that during the injection of melittin, the adsorbed mass is initially underestimated due to the high friction of the protein-solution, see also Braun et al., 2005 [5] for a discussion. During the subsequent buffer injection the correct mass is measured.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2572191&req=5

pone-0003610-g002: Combined mode measurements of vesicle and melittin adsorption on the cantilever sensor.The positive controls were pre-functionalized in such a way that vesicles only bind to the upper cantilever surface and melittin does not bind at all. The lower graph displays the mass adsorption and the upper graph reveals the surface stress development measured simultaneously. Note that the surface stress represents the differential signal between the positively and negatively functionalized cantilevers (two cantilevers each). The experiment was performed in 11 sections: (I) Baseline recording in buffer. (II, VI, VIII) Injections of 500 ng/ml DOPC vesicles. (III, V, VII, IX, XI) Buffer injections. (IV, X) Melittin injections (1 µM). Note that during the injection of melittin, the adsorbed mass is initially underestimated due to the high friction of the protein-solution, see also Braun et al., 2005 [5] for a discussion. During the subsequent buffer injection the correct mass is measured.
Mentions: Supplemental Figure S2 (panel A) depicts the workflow of the main experiments and the results are shown in Figure 2. The adsorption of lipid to the cantilever has to be controlled carefully since asymmetrical functionalization of the cantilevers was crucial for detection of the static cantilever bending (a single sided coating was not a prerequisite for the mass adsorption signal). This was achieved by a specific pre-functionalization of the sensor interfaces.

Bottom Line: Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro.This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology.The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

View Article: PubMed Central - PubMed

Affiliation: National Center of Competence for Research in Nanoscience, Institute of Physics, University of Basel, Basel, Switzerland.

ABSTRACT
Molecular interaction is a key concept in our understanding of the biological mechanisms of life. Two physical properties change when one molecular partner binds to another. Firstly, the masses combine and secondly, the structure of at least one binding partner is altered, mechanically transducing the binding into subsequent biological reactions. Here we present a nanomechanical micro-array technique for bio-medical research, which not only monitors the binding of effector molecules to their target but also the subsequent effect on a biological system in vitro. This label-free and real-time method directly and simultaneously tracks mass and nanomechanical changes at the sensor interface using micro-cantilever technology. To prove the concept we measured lipid vesicle (approximately 748*10(6) Da) adsorption on the sensor interface followed by subsequent binding of the bee venom peptide melittin (2840 Da) to the vesicles. The results show the high dynamic range of the instrument and that measuring the mass and structural changes simultaneously allow a comprehensive discussion of molecular interactions.

Show MeSH
Related in: MedlinePlus