4KQ8

Structure of Recombinant Human Cytochrome P450 Aromatase

Summary for 4KQ8

• Entry DOI: 10.2210/pdb4kq8/pdb
• Related: 3EQM 3S79 3S7S 4GL5 4GL7
• Descriptor: Cytochrome P450 19A1, PROTOPORPHYRIN IX CONTAINING FE, 4-ANDROSTENE-3-17-DIONE, ... (5 entities in total)
• Functional Keywords: aromatase, androgen, estrogen, cytochrome p450, oxidoreductase, estrogen synthetase, cytochrome p450 reductase
• Biological source: Homo sapiens (human)
• Cellular location: Membrane; Peripheral membrane protein: P11511
• Total number of polymer chains: 1
• Total formula weight: 55683.77

Authors

Ghosh, D.,Di Nardo, G.,Griswold, J. (deposition date: 2013-05-14, release date: 2013-08-21, Last modification date: 2024-02-28)

Primary citation

Lo, J.,Di Nardo, G.,Griswold, J.,Egbuta, C.,Jiang, W.,Gilardi, G.,Ghosh, D.
Structural basis for the functional roles of critical residues in human cytochrome p450 aromatase.
Biochemistry, 52:5821-5829, 2013

PubMed Abstract: Cytochrome P450 aromatase (CYP19A1) is the only enzyme known to catalyze the biosynthesis of estrogens from androgens. The crystal structure of human placental aromatase (pArom) has paved the way toward understanding the structure-function relationships of this remarkable enzyme. Using an amino terminus-truncated recombinant human aromatase (rArom) construct, we investigate the roles of key amino acids in the active site, at the intermolecular interface, inside the access channel, and at the lipid-protein boundary for their roles in enzyme function and higher-order organization. Replacing the active site residue D309 with an N yields an inactive enzyme, consistent with its proposed involvement in aromatization. Mutation of R192 at the lipid interface, pivotal to the proton relay network in the access channel, results in the loss of enzyme activity. In addition to the distal catalytic residues, we show that mutation of K440 and Y361 of the heme-proximal region critically interferes with substrate binding, enzyme activity, and heme stability. The D-E loop deletion mutant Del7 that disrupts the intermolecular interaction significantly reduces enzyme activity. However, the less drastic Del4 and point mutants E181A and E181K do not. Furthermore, native gel electrophoresis, size-exclusion chromatography, and analytical ultracentrifugation are used to show that mutations in the intermolecular interface alter the quaternary organization of the enzyme in solution. As a validation for interpretation of the mutational results in the context of the innate molecule, we determine the crystal structure of rArom to show that the active site, tertiary, and quaternary structures are identical to those of pArom.

PubMed: 23899247
DOI: 10.1021/bi400669h

Experimental method

X-RAY DIFFRACTION (3.29 Å)