Monolayers of a model anesthetic-binding membrane protein: formation, characterization, and halothane-binding affinity.
hbAP0 is a model membrane protein designed to possess an anesthetic-binding cavity in its hydrophilic domain and a cation channel in its hydrophobic domain. Grazing incidence x-ray diffraction shows that hbAP0 forms four-helix bundles that are vectorially oriented within Langmuir monolayers at the air-water interface. Single monolayers of hbAP0 on alkylated solid substrates would provide an optimal system for detailed structural and dynamical studies of anesthetic-peptide interaction via x-ray and neutron scattering and polarized spectroscopic techniques. Langmuir-Blodgett and Langmuir-Schaeffer deposition and self-assembly techniques were used to form single monolayer films of the vectorially oriented peptide hbAP0 via both chemisorption and physisorption onto suitably alkylated solid substrates. The films were characterized by ultraviolet absorption, ellipsometry, circular dichroism, and polarized Fourier transform infrared spectroscopy. The alpha-helical secondary structure of the peptide was retained in the films. Under certain conditions, the average orientation of the helical axis was inclined relative to the plane of the substrate, approaching perpendicular in some cases. The halothane-binding affinity of the vectorially oriented hbAP0 peptide in the single monolayers, with the volatile anesthetic introduced into the moist vapor environment of the monolayer, was found to be similar to that for the detergent-solubilized peptide.
Departments of Chemistry and Anesthesiology, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323, USA.
Inna Y
IY
Churbanova
Andrey
A
Tronin
Joseph
J
Strzalka
Thomas
T
Gog
Ivan
I
Kuzmenko
Jonas S
JS
Johansson
J Kent
JK
Blasie
D000327:Adsorption; D000595:Amino Acid Sequence; D000777:Anesthetics / Q000378:metabolism*; D018685:Anesthetics, Inhalation / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D002942:Circular Dichroism; D006221:Halothane / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D013055:Spectrophotometry, Infrared
D000777:Anesthetics; D018685:Anesthetics, Inhalation; D002352:Carrier Proteins; D008565:Membrane Proteins; D006221:Halothane