title: Characterization of the sensor domain of QseE histidine kinase from Escherichia coli.
abstract: In enterohemorrhagic Escherichia coli (EHEC), the QseEF two-component system causes attaching and effacing (AE) lesion on epithelial cells. QseE histidine kinase senses the host hormone epinephrine, sulfate, and phosphate; it also regulates QseF response regulator, which activates LEE gene that encodes AE lesion. In order to understand the recognition of ligand molecules and signal transfer mechanism in pathogenic bacteria, structural studies of the sensor domain of QseE of Escherichia coli should be conducted. In this study, we describe the overexpression, purification, and structural and biophysical properties of the sensor domain of QseE. The fusion protein had a 6×His tag at its N-terminus; this protein was overexpressed as inclusion bodies in E. coli BL21 (DE3). The protein was denatured in 7M guanidine hydrochloride and refolded by dialysis. The purification of the refolded protein was carried out using Ni-NTA affinity column and size-exclusion chromatography. Thereafter, the characteristics of the refolded protein were determined from NMR, CD, and MALS spectroscopies. In a pH range of 7.4-5.0, the folded protein existed in a monomeric form with a predominantly helical structure. (1)H-(15)N HSQC NMR spectra shows that approximately 93% backbone amide peaks are detected at pH 5.0, suggesting that the number of backbone signals is sufficient for NMR studies. These data might provide an opportunity for structural and functional studies of the sensor domain of QseE.
authors.affiliation: Protein Structure Group, Korea Basic Science Institute (KBSI), 162 Yeongudanji-Ro, Ochang, Chungbuk, 28119, Republic of Korea; College of Pharmacy, Korea University, 2511 Sejong-Ro, Sejong, 30019, Republic of Korea.
authors.forename: Kwon Joo
authors.initials: KJ
authors.lastname: Yeo
authors.affiliation: Protein Structure Group, Korea Basic Science Institute (KBSI), 162 Yeongudanji-Ro, Ochang, Chungbuk, 28119, Republic of Korea.
authors.forename: Jin-Wan
authors.initials: JW
authors.lastname: Park
authors.affiliation: Protein Structure Group, Korea Basic Science Institute (KBSI), 162 Yeongudanji-Ro, Ochang, Chungbuk, 28119, Republic of Korea.
authors.forename: Eun-Hee
authors.initials: EH
authors.lastname: Kim
authors.affiliation: College of Pharmacy, Korea University, 2511 Sejong-Ro, Sejong, 30019, Republic of Korea.
authors.forename: Young Ho
authors.initials: YH
authors.lastname: Jeon
authors.affiliation: Division of Biotechnology, College of Life Sciences & Biotechnology, Korea University, Seoul, 02841, Republic of Korea. Electronic address: chahong@korea.ac.kr.
authors.forename: Kwang Yeon
authors.initials: KY
authors.lastname: Hwang
authors.affiliation: Protein Structure Group, Korea Basic Science Institute (KBSI), 162 Yeongudanji-Ro, Ochang, Chungbuk, 28119, Republic of Korea. Electronic address: haekap@kbsi.re.kr.
authors.forename: Hae-Kap
authors.initials: HK
authors.lastname: Cheong
mesh_terms: D054324:Enterohemorrhagic Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D019906:Nuclear Magnetic Resonance, Biomolecular; D000072417:Protein Domains; D058849:Protein Refolding; D017433:Protein Structure, Secondary; D011941:Receptors, Adrenergic / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
chemical_list: D029968:Escherichia coli Proteins; C540351:QseE protein, E coli; D011941:Receptors, Adrenergic; D011993:Recombinant Fusion Proteins
keywords: Histidine kinase; QseE; QseE sensor domain; Quorum sensing; Refolding