PDBsum entry 1e9x

Oxidoreductase

PDB id: 1e9x

Contents

Protein chain

449 a.a. *

Ligands

HEM-PIM

Waters ×282

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Crystal structure of cytochrome p450 14alpha -Sterol demethylase (cyp51) from mycobacterium tuberculosis in complex with azole inhibitors.

Authors

L.M.Podust, T.L.Poulos, M.R.Waterman.

Ref.

Proc Natl Acad Sci U S A, 2001, 98, 3068-3073. [DOI no: 10.1073/pnas.061562898]

PubMed id

11248033

Abstract

Cytochrome P450 14alpha-sterol demethylases (CYP51) are essential enzymes in sterol biosynthesis in eukaryotes. CYP51 removes the 14alpha-methyl group from sterol precursors such as lanosterol, obtusifoliol, dihydrolanosterol, and 24(28)-methylene-24,25-dihydrolanosterol. Inhibitors of CYP51 include triazole antifungal agents fluconazole and itraconazole, drugs used in treatment of topical and systemic mycoses. The 2.1- and 2.2-A crystal structures reported here for 4-phenylimidazole- and fluconazole-bound CYP51 from Mycobacterium tuberculosis (MTCYP51) are the first structures of an authentic P450 drug target. MTCYP51 exhibits the P450 fold with the exception of two striking differences-a bent I helix and an open conformation of BC loop-that define an active site-access channel running along the heme plane perpendicular to the direction observed for the substrate entry in P450BM3. Although a channel analogous to that in P450BM3 is evident also in MTCYP51, it is not open at the surface. The presence of two different channels, with one being open to the surface, suggests the possibility of conformationally regulated substrate-in/product-out openings in CYP51. Mapping mutations identified in Candida albicans azole-resistant isolates indicates that azole resistance in fungi develops in protein regions involved in orchestrating passage of CYP51 through different conformational stages along the catalytic cycle rather than in residues directly contacting fluconazole. These new structures provide a basis for rational design of new, more efficacious antifungal agents as well as insight into the molecular mechanism of P450 catalysis.

Figure 1.
Fig. 1. Ribbon representation of the MTCYP51 structures with the inhibitors bound. Front (A) and top (B) views of the of 4-PI- (yellow) and FLU- (blue) bound MTCYP51 superimposed with an rms deviation of 0.45 Å. Superimpositions for all figures were done by using two-step fitting as implemented in SWISS-PDB VIEWER (33). The first step was performed by using entire structures; for the second step, an rms-deviation cutoff of 1.8 Å was used to select the most structurally homologous regions for subsequent fitting. The second round results in better fitting of the most homologous regions and further divergence of less homologous regions. Heme, red; 4-PI, orange; FLU, light-blue. The I helix is shown also in red. A large cavity of 2,600 Å3, shown in blue, leads from the active site to the molecular surface along the protein domain interface (channel 2). Structural elements significantly deviating among P450 structures are labeled in black, and -sheets that are part of the putative substrate-binding site are labeled in red. All figures, if not otherwise indicated, are generated by using SWISS-PDB VIEWER (33).

Figure 2.
Fig. 2. Superimposition and alignment of the I helix in known P450 structures. Front (A) and top (B) views of the I helix from superimposed P450 structures assigned in sequence-alignment shown in C. Each structure was superimposed pairwise with MTCYP51 so that rms deviation for the most structurally homologous regions did not exceed 1.2 Å. (C) Alignment of the I helix sequences performed by using BCM SEARCH LAUNCHER (34). Residues identical or homologous in at least half of the compared sequences are shaded in dark or light, respectively. The position of conserved glycine is marked according to MTCYP51 sequence (P77901).