of Chemistry, 2001, 4, 6 [ISSN: 1099-8292].
Substrate access to cytochrome P450cam investigated
by molecular dynamics simulations:
An interactive look at the underlying mechanisms
Susanna K. Ldemann,1 Razif R. Gabdoulline,
2 Valre Lounnas,3 and Rebecca C. Wade2
European Molecular Biology Laboratory, Meyerhofstr.1, 69117 Heidelberg,
(1) Bioinformatics Unit, Research Institute of Molecular Pathology, Dr.
Bohrg.7, 103 0 Wien, Austria.
(2) European Media Laboratory, Villa Bosch, Schloss-Wolfsbrunnenweg 33,
D-69118 Heidelberg and European Molecular Biology Laboratory Heidelberg, Meyerhofstr
(3) Molecular Design and Informatics, N.V. Organon, Molenstraat 110, PO
Box 20, 5340 BH Oss, The Netherlands.
Publication date: May 30, 2001 14:31:00 GMT
cytochrome P450, molecular dynamics simulation, protein-ligand
binding, protein dynamics, molecular animation, random expulsion molecular
dynamics, interactive molecular graphics visualisation
Cytochrome P450cam (P450cam) from Pseudomonas putida has long provided
a paradigm for structural understanding of cytochrome P450s, a ubiquitous
protein family with functions including the synthesis and degradation of
physiologically important compounds, e.g. steroids and prostaglandins, and
of many xenobiotics, e.g. drugs and procarcinogens. The mechanism by which
camphor, the natural substrate of P450cam, accesses the buried active site
is not clear. While there is recent crystallographic and simulation evidence
for opening of a substrate access channel in cytochrome P450BM-3 (P450BM-3),
for P450cam such conformational changes upon substrate access have not been
observed either in different crystal structures or by standard molecular
dynamics simulations. We therefore developed a new simulation technique,
the random expulsion molecular dynamics method, for probing ligand exit from
buried active sites by imposing an artificial randomly oriented force on
the ligand in addition to the standard molecular dynamics force field (Ldemann,
S. K., Lounnas, V. and Wade, R. C., J. Mol. Biol., 303, 797-811, 2000). In
the present paper, representative animations for the three exit pathways
obtained by the random expulsion molecular dynamics method are shown. The
method was tested in simulations of the substrate-bound structure of P450BM-3,
for which an animation is also shown. The protein dynamics involved in specific
substrate exit mechanisms, in particular the transient fluctuations and perturbation
of salt-links can be inspected directly in an animation and thus give much
better insight into possible access/exit mechanisms than static figures.
In addition, center of mass traces of the substrates along different expulsion
pathways in P450cam can be examined interactively using MolSurfer, a java-based
protein structure viewer originally employed for navigating molecular interfaces
(Gabdoulline, R. R., Wade, R. C. and Walther, D., Trends Biochem. Sci 24,
285-7 (1999)). In the present paper, this tool is applied for navigating along
the ligand exit pathways in cytochrome P450cam, thereby permitting interactive
viewing of the local protein environment simultaneously with precomputed
dynamic parameters from molecular dynamics simulations and experimentally
determined crystallographic temperature factors.