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Structure paper

タイトルCrystal structure of the flavin reductase of Acinetobacter baumannii p-hydroxyphenylacetate 3-hydroxylase (HPAH) and identification of amino acid residues underlying its regulation by aromatic ligands.
ジャーナル・号・ページArch Biochem Biophys, Vol. 653, Page 24-38, Year 2018
掲載日2018年9月1日
著者Anan Yuenyao / Nopphon Petchyam / Nuntaporn Kamonsutthipaijit / Pimchai Chaiyen / Danaya Pakotiprapha /
PubMed 要旨The first step in the degradation of p-hydroxyphenylacetic acid (HPA) is catalyzed by the two-component enzyme p-hydroxyphenylacetate 3-hydroxylase (HPAH). The two components of Acinetobacter ...The first step in the degradation of p-hydroxyphenylacetic acid (HPA) is catalyzed by the two-component enzyme p-hydroxyphenylacetate 3-hydroxylase (HPAH). The two components of Acinetobacter baumannii HPAH are known as C and C, respectively. C is a flavin reductase that uses NADH to generate reduced flavin mononucleotide (FMNH), which is used by C in the hydroxylation of HPA. Interestingly, although HPA is not directly involved in the reaction catalyzed by C, the presence of HPA dramatically increases the FMN reduction rate. Amino acid sequence analysis revealed that C contains two domains: an N-terminal flavin reductase domain, and a C-terminal MarR domain. Although MarR proteins typically function as transcription regulators, the MarR domain of C was found to play an auto-inhibitory role. Here, we report a crystal structure of C and small-angle X-ray scattering (SAXS) studies that revealed that C undergoes a substantial conformational change in the presence of HPA, concomitant with the increase in the rate of flavin reduction. Amino acid residues that are important for HPA binding and regulation of C activity were identified by site-directed mutagenesis. Amino acid sequence similarity analysis revealed several as yet uncharacterized flavin reductases with N- or C-terminal fusions.
リンクArch Biochem Biophys / PubMed:29940152
手法SAS (X-ray synchrotron) / X線回折
解像度2.898 Å
構造データ

SASDD24: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of H170A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD28: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD34: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of H170A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD38: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD44: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of H170A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD48: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD54: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of H170A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD58: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E251A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD64: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of H170A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD68: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E251A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD74: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of S172A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD78: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E251A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD84: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of S172A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDD94: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of S172A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDA4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of S172A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDB4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of S172A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDC4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of S172A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDD4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of N174A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDE4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of N174A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDF4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of N174A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDG4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of N174A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDH4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of N174A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDJ4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of Y207A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDK4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of Y207A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDL4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of Y207A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDM4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of Y207A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDN4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of Y207A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDN7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E248A/E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDP4: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of Y207A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDP7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E248A/E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDQ7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E248A/E251A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDR7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E248A/E251A C1 in the presence of 1 mM p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDS7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E248A/E251A C1 in the presence of 1 mM p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDT3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 2 mg/ml of wild-type C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDT7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E248A/E251A C1 in the presence of 1 mM p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDU3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 4 mg/ml of Wild-type C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDU7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E248A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDV3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 8 mg/ml of Wild-type C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDV7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E248A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDW3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 2 mg/ml of Wild-type C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDW7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E248A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDX3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 4 mg/ml of Wild-type C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDX7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of E248A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDY3: p-hydroxyphenylacetate 3-hydroxylase, reductase component: 8 mg/ml of Wild-type C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDY7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 4 mg/ml of E248A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDZ3: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 2 mg/ml of H170A C1 in the absence of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

SASDDZ7: p-hydroxyphenylacetate 3-hydroxylase, reductase component (mutant): 8 mg/ml of E248A C1 in 1 mM of p-hydroxyphenylacetic acid (HPA)
手法: SAXS/SANS

PDB-5zc2:
Acinetobacter baumannii p-hydroxyphenylacetate 3-hydroxylase (HPAH), reductase component (C1)
手法: X-RAY DIFFRACTION / 解像度: 2.898 Å

化合物

ChemComp-FMN:
FLAVIN MONONUCLEOTIDE / FMN

由来
  • acinetobacter baumannii (バクテリア)
キーワードFLAVOPROTEIN / Flavin reductase / MarR

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