2YOO

Cholest-4-en-3-one bound structure of CYP142 from Mycobacterium smegmatis

PDB DOI: https://doi.org/10.2210/pdb2YOO/pdb

Classification: OXIDOREDUCTASE
Organism(s): Mycolicibacterium smegmatis MC2 155
Expression System: Escherichia coli
Mutation(s): No

Deposited: 2012-10-25 Released: 2013-02-27
Deposition Author(s): Garcia-Fernandez, E., Frank, D.J., Galan, B., Kells, P.M., Podust, L.M., Garcia, J.L., Ortiz de Montellano, P.R.

Experimental Data Snapshot

Method: X-RAY DIFFRACTION
Resolution: 1.69 Å
R-Value Free: 0.219
R-Value Work: 0.165
R-Value Observed: 0.168

wwPDB Validation 3D Report Full Report

Ligand Structure Quality Assessment

This is version 1.4 of the entry. See complete history.

Literature

A Highly Conserved Mycobacterial Cholesterol Catabolic Pathway.

Garcia-Fernandez, E., Frank, D.J., Galan, B., Kells, P.M., Podust, L.M., Garcia, J.L., Ortiz de Montellano, P.R.

(2013) Environ Microbiol 15: 2342

PubMed: 23489718 Search on PubMedSearch on PubMed Central
DOI: https://doi.org/10.1111/1462-2920.12108
Primary Citation of Related Structures:
2YOO, 4APY

PubMed Abstract:
Degradation of the cholesterol side-chain in Mycobacterium tuberculosis is initiated by two cytochromes P450, CYP125A1 and CYP142A1, that sequentially oxidize C26 to the alcohol, aldehyde and acid metabolites. Here we report characterization of the homologous enzymes CYP125A3 and CYP142A2 from Mycobacterium smegmatis mc(2) 155. Heterologously expressed, purified CYP125A3 and CYP142A2 bound cholesterol, 4-cholesten-3-one, and antifungal azole drugs. CYP125A3 or CYP142A2 reconstituted with spinach ferredoxin and ferredoxin reductase efficiently hydroxylated 4-cholesten-3-one to the C-26 alcohol and subsequently to the acid. The X-ray structures of both substrate-free CYP125A3 and CYP142A2 and of cholest-4-en-3-one-bound CYP142A2 reveal significant differences in the substrate binding sites compared with the homologous M. tuberculosis proteins. Deletion only of cyp125A3 causes a reduction of both the alcohol and acid metabolites and a strong induction of cyp142 at the mRNA and protein levels, indicating that CYP142A2 serves as a functionally redundant back up enzyme for CYP125A3. In contrast to M. tuberculosis, the M. smegmatis Δcyp125Δcyp142 double mutant retains its ability to grow on cholesterol albeit with a diminished capacity, indicating an additional level of redundancy within its genome.

Organizational Affiliation

Department of Environmental Biology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain.

Macromolecule Content

Total Structure Weight: 187.15 kDa
Atom Count: 14,507
Modelled Residue Count: 1,580
Deposited Residue Count: 1,628
Unique protein chains: 1

Macromolecules

Find similar proteins by:

(by identity cutoff) | 3D Structure

Entity ID: 1
Molecule	Chains	Sequence Length	Organism	Details	Image
P450 HEME-THIOLATE PROTEIN	A, B, C, D	407	Mycolicibacterium smegmatis MC2 155	Mutation(s): 0
UniProt
Find proteins for A0R4Q6 (Mycolicibacterium smegmatis (strain ATCC 700084 / mc(2)155)) Explore A0R4Q6 Go to UniProtKB: A0R4Q6
Entity Groups
Sequence Clusters	30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt Group	A0R4Q6
Sequence Annotations Expand
Reference Sequence

Small Molecules

Ligands 3 Unique
ID	Chains	Name / Formula / InChI Key	2D Diagram	3D Interactions
HEM Query on HEM Download Ideal Coordinates CCD File SDF format, chain E [auth A] SDF format, chain G [auth B] SDF format, chain J [auth C] SDF format, chain L [auth D] MOL2 format, chain E [auth A] MOL2 format, chain G [auth B] MOL2 format, chain J [auth C] MOL2 format, chain L [auth D]	E [auth A], G [auth B], J [auth C], L [auth D]	PROTOPORPHYRIN IX CONTAINING FE C₃₄ H₃₂ Fe N₄ O₄ KABFMIBPWCXCRK-RGGAHWMASA-L		Interactions Focus chain E [auth A] Focus chain G [auth B] Focus chain J [auth C] Focus chain L [auth D] Interactions & Density Focus chain E [auth A] Focus chain G [auth B] Focus chain J [auth C] Focus chain L [auth D]
K2B Query on K2B Download Ideal Coordinates CCD File SDF format, chain F [auth A] SDF format, chain H [auth B] SDF format, chain K [auth C] SDF format, chain M [auth D] MOL2 format, chain F [auth A] MOL2 format, chain H [auth B] MOL2 format, chain K [auth C] MOL2 format, chain M [auth D]	F [auth A], H [auth B], K [auth C], M [auth D]	(8ALPHA,9BETA)-CHOLEST-4-EN-3-ONE C₂₇ H₄₄ O NYOXRYYXRWJDKP-GYKMGIIDSA-N		Interactions Focus chain F [auth A] Focus chain H [auth B] Focus chain K [auth C] Focus chain M [auth D] Interactions & Density Focus chain F [auth A] Focus chain H [auth B] Focus chain K [auth C] Focus chain M [auth D]
MG Query on MG Download Ideal Coordinates CCD File SDF format, chain I [auth B] MOL2 format, chain I [auth B]	I [auth B]	MAGNESIUM ION Mg JLVVSXFLKOJNIY-UHFFFAOYSA-N		Interactions Focus chain I [auth B] Interactions & Density Focus chain I [auth B]

Binding Affinity Annotations
ID	Source	Binding Affinity
K2B	Binding MOAD: 2YOO	Kd: 520 (nM) from 1 assay(s)

Experimental Data & Validation

Experimental Data

Method: X-RAY DIFFRACTION
Resolution: 1.69 Å
R-Value Free: 0.219
R-Value Work: 0.165
R-Value Observed: 0.168
Space Group: P 1 2₁ 1

Unit Cell:

Length ( Å )	Angle ( ˚ )
a = 56.683	α = 90
b = 106.213	β = 90.67
c = 126.536	γ = 90

Software Package:

Software Name	Purpose
REFMAC	refinement
MOSFLM	data reduction
SCALA	data scaling
MOLREP	phasing

Structure Validation

View Full Validation Report

Ligand Structure Quality Assessment

Entry History

Deposition Data

Released Date: 2013-02-27

Deposition Author(s):

Ortiz de Montellano, P.R.

Revision History (Full details and data files)

Version 1.0: 2013-02-27
Type: Initial release
Version 1.1: 2013-05-01
Changes: Database references
Version 1.2: 2013-08-14
Changes: Database references
Version 1.3: 2018-02-07
Changes: Database references, Source and taxonomy
Version 1.4: 2023-12-20
Changes: Data collection, Database references, Derived calculations, Other, Refinement description