Structure of the quinoline N-hydroxylating cytochrome P450 RauA, an essential enzyme that confers antibiotic activity on aurachin alkaloids

Yoshiaki Yasutake; Wataru Kitagawa; Miyako Hata; Taiki Nishioka; Taro Ozaki; Makoto Nishiyama; Tomohisa Kuzuyama; Tomohiro Tamura

doi:10.1016/j.febslet.2013.11.016

Structure of the quinoline N-hydroxylating cytochrome P450 RauA, an essential enzyme that confers antibiotic activity on aurachin alkaloids

FEBS Lett. 2014 Jan 3;588(1):105-10. doi: 10.1016/j.febslet.2013.11.016. Epub 2013 Nov 20.

Authors

Yoshiaki Yasutake¹, Wataru Kitagawa², Miyako Hata³, Taiki Nishioka⁴, Taro Ozaki⁵, Makoto Nishiyama⁵, Tomohisa Kuzuyama⁵, Tomohiro Tamura²

Affiliations

¹ Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 2-17-2-1 Tsukisamu-Higashi, Toyohira-ku, Sapporo 062-8517, Japan. Electronic address: [email protected].
² Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 2-17-2-1 Tsukisamu-Higashi, Toyohira-ku, Sapporo 062-8517, Japan; Graduate School of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kita-ku, Sapporo 060-8589, Japan.
³ Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 2-17-2-1 Tsukisamu-Higashi, Toyohira-ku, Sapporo 062-8517, Japan.
⁴ Graduate School of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kita-ku, Sapporo 060-8589, Japan.
⁵ Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.

PMID: 24269679
DOI: 10.1016/j.febslet.2013.11.016

Abstract

The cytochrome P450 RauA from Rhodococcus erythropolis JCM 6824 catalyzes the hydroxylation of a nitrogen atom in the quinolone ring of aurachin, thereby conferring strong antibiotic activity on the aurachin alkaloid. Here, we report the crystal structure of RauA in complex with its substrate, a biosynthetic intermediate of aurachin RE. Clear electron density showed that the quinolone ring is oriented parallel to the porphyrin plane of the heme cofactor, while the farnesyl chain curls into a U-shape topology and is buried inside the solvent-inaccessible hydrophobic interior of RauA. The nearest atom from the heme iron is the quinolone nitrogen (4.3Å), which is consistent with RauA catalyzing the N-hydroxylation of the quinolone ring to produce mature aurachin RE.

Keywords: Antibiotic; Aurachin; CYP; Cytochrome P450; HPLC; Menaquinone; RMSD; Rhodococcus erythropolis; cytochrome P450 monooxygenase; high-performance liquid chromatography; root–mean–square deviation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Alkaloids / chemistry
Alkaloids / metabolism
Amino Acid Sequence
Bacterial Proteins / chemistry*
Bacterial Proteins / genetics
Bacterial Proteins / metabolism
Biocatalysis
Crystallography, X-Ray
Cytochrome P-450 Enzyme System / chemistry*
Cytochrome P-450 Enzyme System / genetics
Cytochrome P-450 Enzyme System / metabolism
Genes, Essential
Hydroxylamines / chemistry*
Hydroxylamines / metabolism
Hydroxylation
Kinetics
Models, Molecular
Molecular Sequence Data
Molecular Structure
Phylogeny
Protein Binding
Protein Structure, Tertiary
Quinolines / chemistry*
Quinolines / metabolism
Quinolones / chemistry*
Quinolones / metabolism
Rhodococcus / enzymology
Rhodococcus / genetics
Rhodococcus / metabolism
Sequence Homology, Amino Acid
Spectrometry, Fluorescence
Substrate Specificity

Substances

Alkaloids
Bacterial Proteins
Hydroxylamines
Quinolines
Quinolones
RauA protein, Rhodococcus erythropolis
aurachin RE
Cytochrome P-450 Enzyme System
quinoline