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          Institute: MPI für Biochemie     Collection: Structural Biology (W. Baumeister)     Display Documents



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ID: 318465.0, MPI für Biochemie / Structural Biology (W. Baumeister)
Molecular recognition of histidine-tagged molecules by metal-chelating lipids monitored by fluorescence energy transfer and correlation spectroscopy
Authors:Dorn, I. T.; Neumaier, K. R.; Tampe, R.
Date of Publication (YYYY-MM-DD):1998-04-01
Title of Journal:Journal of the American Chemical Society
Volume:120
Issue / Number:12
Start Page:2753
End Page:2763
Review Status:not specified
Audience:Not Specified
Abstract / Description:Complex binding of proteins by metal-chelating Lipids via surface-exposed or protein-engineered histidines provides an universal and powerful concept for the orientation and two-dimensional crystallization of proteins at self-organized interfaces. To demonstrate pair formation between individual histidine-tagged molecules and chelator lipids on the molecular level, we have synthesized novel lipids bearing both a Ni-NTA chelator and a fluorescent group. These lipids serve as spectroscopic probes to visualize directly the molecular recognition of fluorescence-labeled histidine-tagged peptides by metal-chelating lipids using fluorescence resonance energy transfer (FRET). The molecular docking to chelator lipids assembled in mono- or bilayers is highly specific, revealing only 3% unspecific adsorption and a binding constant of 3 mu M. The affinity constant was confirmed by fluorescence correlation spectroscopy (FCS) on single molecules, where the ratio of lipid-bound to free was analyzed by their intrinsically different diffusion times passing through a confocal volume of about 1 fL. By using a model peptide most of the electrostatic and steric contribution to the binding process can be neglected. Therefore, the affinity constant can serve as a standard value for the binding of histidine-tagged proteins to chelator lipid interfaces. [References: 51]
Free Keywords:Surface-plasmon resonance; Air-water-interface; Reversible immobilization; Recombinant proteins; Membranes; Monolayers; Binding; Crystallization; Coordination; Bilayers.; Chemistry. Chemistry & analysis.
External Publication Status:published
Document Type:Article
Communicated by:Anton Hillebrand
Affiliations:MPI für Biochemie/Structural Biology (W. Baumeister)
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