Applications of Biomembranes in Drug Discovery

Ye, Fang; Hong, Yulong; Webb, Brian; Lahiri, Joydeep

doi:10.1557/mrs2006.140

Cited by 24 publications

(15 citation statements)

References 52 publications

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“…The MST results are in good agreement with previous SPR data showing a dissociation constant of 1–2 nM for the binding of neurotensin to an NTS1 fusion construct, [42] while slightly lower affinities have previously been found for a fluorescently labeled (TAMRA) neurotensin derivative by fluorescence correlation spectroscopy (K D =7±3 nM, Harding, unpublished results), although other fluorophores did not affect the affinity (K D =1.4 nM for Cy5-neurotensin). [43] K D dependence on the choice of fluorophores has previously been observed for other fluorescent ligand derivatives as well. [44] We also studied binding of a known inverse agonist, SR48692, to NTS1B and obtained a K D of 15±11 nM using label-free MST (Fig.…”

Section: Resultsmentioning

confidence: 74%

Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions

Seidel

Dijkman²,

Lea³

et al. 2013

Methods

541

478

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Microscale thermophoresis (MST) allows for quantitative analysis of protein interactions in free solution and with low sample consumption. The technique is based on thermophoresis, the directed motion of molecules in temperature gradients. Thermophoresis is highly sensitive to all types of binding-induced changes of molecular properties, be it in size, charge, hydration shell or conformation. In an all-optical approach, an infrared laser is used for local heating, and molecule mobility in the temperature gradient is analyzed via fluorescence. In standard MST one binding partner is fluorescently labeled. However, MST can also be performed label-free by exploiting intrinsic protein UV-fluorescence. Despite the high molecular weight ratio, the interaction of small molecules and peptides with proteins is readily accessible by MST. Furthermore, MST assays are highly adaptable to fit to the diverse requirements of different biomolecules, such as membrane proteins to be stabilized in solution. The type of buffer and additives can be chosen freely. Measuring is even possible in complex bioliquids like cell lysate allowing close to in vivo conditions without sample purification. Binding modes that are quantifiable via MST include dimerization, cooperativity and competition. Thus, its flexibility in assay design qualifies MST for analysis of biomolecular interactions in complex experimental settings, which we herein demonstrate by addressing typically challenging types of binding events from various fields of life science.

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Section: Resultsmentioning

confidence: 74%

Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions

Seidel

Dijkman²,

Lea³

et al. 2013

Methods

541

478

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“…The lipid-protein interactions may lead to phase transitions at hydrocarbon chains of lipid molecules, change the thickness of the membrane, the water content in the membrane or the net charge in the polar head group region. Moreover, both macroscopic and microscopic properties of a biological membrane such as permeability to ions, capacitance, lateral composition, concentration, orientation and physical state of lipid molecules and the activity and structure of associated proteins can be affected by drugs, changes in electrolyte composition and concentration or the static electric field [144,145]. A combination of IRS, sensitive to the structure of lipid and proteins molecules, to electrochemistry testing macroscopic properties of model membranes such as redox activity, membrane capacity or permeability to ions, giving a possibility to monitor simultaneously potential-dependent changes in lipid bilayers arising from the lipid-protein interaction [39][40][41][42]146] or redox activity of a protein embedded in the membrane [147][148][149][150].…”

Section: Application Of In Situ Pm Irras For the Determination Of Thementioning

confidence: 99%

Application of Polarization Modulation Infrared Reflection Absorption Spectrosocpy Under Electrochemical Control for Structural Studies of Biomimetic Assemblies

Brand

2015

Zeitschrift Für Physikalische Chemie

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An ideal bioanalytical method would allow carrying out sensitive, selective, reproducible, fast, and in situ chemical measurements of the composition, structure and function of complex biological systems. Physical chemistry possesses advanced structure analyzing techniques which are suitable to solve, at a sub-molecular level, the structure of biomolecules. However, the prerequisite of the presence of the electrolyte solution limits the number of applicable structure analyzing techniques. Infrared spectroscopy is one of the most powerful analytical techniques used to identify the structure of biomolecules, monitor structural changes under various experimental environments and physical states. In addition, infrared reflection absorption spectroscopy (IRRAS) has been successfully applied to study supramolecular films at the solid|liquid interface. Assemblies of biomolecules belong to a particularly demanding because they bring specific for the maintenance of their functions experimental requirements which have to be taken into account planning in situ spectroelectrochemical experiments. In this paper potnential of in situ IRRAS under electrochemical control for studies of structural changes in assemblies of biomolecules such as lipid bilayers and protein films is described. Studies of different classes of biomolecules bring certain experimetnal conditions concerning the electrolyte composition, pH value, temperature or chemical nature of the solid support, which have to be fulfilled. Obviously, these conditons may significantly restrict the applicability of the in situ PM IRRAS. The selection of the solid substrate with required optical and electrical properties, the optical window and the electrolyte composition have to be adapted to the needs of biomolecules. Adjustement of experimetnal conditions as well as possibilites of the enlargement of the in situ PM IRRAS for studies of biomimetic assemblies is reviewed in this paper. Furthermore, experimetnal resutls reporting potenial driven changes in models of cell membranes and protein films are reviewed. Pioneering studies aiminig at the determination of structural changes in lipid membranes exposed to physiological electric fields and interacting simultaneously with protiens are described.

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“…Fourth, the characterization of kinetics and affinities of receptor-ligand binding in supported membranes is gaining considerable interest in assaying interactions of pharmacological importance and in the design of membrane-based biosensors (45)(46)(47)(48)(49)(50). The readout in these measurements typically relies on the use of fluorescence-, SPR-, or QCM-based transduction.…”

Section: Introductionmentioning

confidence: 99%

Characterization of Physical Properties of Supported Phospholipid Membranes Using Imaging Ellipsometry at Optical Wavelengths

Howland

Szmodis

Sanii

et al. 2007

Biophysical Journal

113

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Subnanometer-scale vertical z-resolution coupled with large lateral area imaging, label-free, noncontact, and in situ advantages make the technique of optical imaging ellipsometry (IE) highly suitable for quantitative characterization of lipid bilayers supported on oxide substrates and submerged in aqueous phases. This article demonstrates the versatility of IE in quantitative characterization of structural and functional properties of supported phospholipid membranes using previously well-characterized examples. These include 1), a single-step determination of bilayer thickness to 0.2 nm accuracy and large-area lateral uniformity using photochemically patterned single 1,2-dimyristoyl-sn-glycero-3-phosphocholine bilayers; 2), hydration-induced spreading kinetics of single-fluid 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine bilayers to illustrate the in situ capability and image acquisition speed; 3), a large-area morphological characterization of phase-separating binary mixtures of 1,2-dilauroyl-sn-glycero-3-phosphocholine and galactosylceramide; and 4), binding of cholera-toxin B subunits to GM1-incorporating bilayers. Additional insights derived from these ellipsometric measurements are also discussed for each of these applications. Agreement with previous studies confirms that IE provides a simple and convenient tool for a routine, quantitative characterization of these membrane properties. Our results also suggest that IE complements more widely used fluorescence and scanning probe microscopies by combining large-area measurements with high vertical resolution without the use of labeled lipids.

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Applications of Biomembranes in Drug Discovery

Cited by 24 publications

References 52 publications

Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions

Microscale thermophoresis quantifies biomolecular interactions under previously challenging conditions

Application of Polarization Modulation Infrared Reflection Absorption Spectrosocpy Under Electrochemical Control for Structural Studies of Biomimetic Assemblies

Characterization of Physical Properties of Supported Phospholipid Membranes Using Imaging Ellipsometry at Optical Wavelengths

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