Javier L. Baylon, Ivan L. Lenov, Stephen G. Sligar, and Emad Tajkhorshid.
Characterizing the membrane-bound state of cytochrome P450 3A4:
Structure, depth of insertion, and orientation.
Journal of the American Chemical Society, 135:8542-8551, 2013.
(PMC: PMC3682445)
BAYL2013-ET
Cytochrome P450 3A4 (CYP3A4) is the most abundant membrane-associated isoform of
the P450 family in humans and is responsible for biotransformation of more than 50% of
drugs metabolized in the body. Despite the large number of crystallographic structures
available for CYP3A4, no structural information for its membrane-bound state at an atomic
level is available. In order to characterize binding, depth of insertion, membrane
orientation, and lipid interactions of CYP3A4, we have employed a combined experimental
and simulation approach in this study. Taking advantage of a novel membrane
representation, highly mobile membrane mimetic (HMMM), with enhanced lipid mobility
and dynamics, we have been able to capture spontaneous binding and insertion of the
globular domain of the enzyme into the membrane in multiple independent, unbiased
simulations. Despite different initial orientations and positions of the protein in solution,
all the simulations converged into the same membrane-bound configuration with regard to
both the depth of membrane insertion and the orientation of the enzyme on the surface of
the membrane. In tandem, linear dichroism measurements performed on CYP3A4 bound to
Nanodisc membranes were used to characterize the orientation of the enzyme in its
membrane-bound form experimentally. The heme tilt angles measured experimentally are
in close agreement with those calculated for the membrane-bound structures resulted
from the simulations, thereby verifying the validity of the developed model. Membrane
binding of the globular domain in CYP3A4, which appears to be independent of the
presence of the transmembrane helix of the full-length enzyme, significantly reshapes the
protein at the membrane interface, causing conformational changes relevant to access
tunnels leading to the active site of the enzyme.
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