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Published on: December 17, 2013
Structural insights into Escherichia coli polymyxin B resistance protein D with X-ray crystallography and small-angle
Hunho Jo1, Eui Young Jeong2, Jinseong Jeon3
1Department of Chemistry, Pohang University of Science and Technology, 77, Cheongam-Ro, Nam-Gu, Pohang, Gyeongbuk, 790-784, South Korea. jhhsst@postech.ac.kr.
Background:
Polymyxin B resistance protein D (PmrD) plays a key role in the polymyxin B-resistance pathway, as it is the signaling protein that can act as a specific connecter between PmrA/PmrB and PhoP/PhoQ. We conducted structural analysis to characterize Escherichia coli (E. coli) PmrD, which exhibits different features compared with PmrD in other bacteria.
Results:
The X-ray crystal structure of E. coli PmrD was determined at a 2.00 Å resolution, revealing novel information such as the unambiguous secondary structures of the protein and the presence of a disulfide bond. Furthermore, various assays such as native gel electrophoresis, surface plasmon resonance (SPR), size-exclusion chromatography, dynamic light scattering (DLS), and small-angle X-ray scattering (SAXS) measurements, were performed to elucidate the structural and functional role of the internal disulfide bond in E. coli PmrD.
Conclusions:
The structural characteristics of E. coli PmrD were clearly identified via diverse techniques. The findings help explain the different protective mechanism of E. coli compared to other Gram-negative bacteria.
Insights
Structural analysis of Escherichia coli Polymyxin B resistance protein D (PmrD) revealed unique features, including a disulfide bond, aiding in understanding polymyxin resistance mechanisms in Gram-negative bacteria.
Area of Science:
- Microbiology
- Structural Biology
- Biochemistry
Background:
- Polymyxin B resistance protein D (PmrD) is crucial for polymyxin B resistance.
- PmrD acts as a signaling protein connecting PmrA/PmrB and PhoP/PhoQ pathways.
- Escherichia coli PmrD displays distinct characteristics compared to PmrD in other bacteria.
Purpose of the Study:
- To characterize the structure of Escherichia coli PmrD.
- To elucidate the structural and functional role of the internal disulfide bond in E. coli PmrD.
Main Methods:
- X-ray crystallography (2.00 Å resolution)
- Native gel electrophoresis
- Surface plasmon resonance (SPR)
- Size-exclusion chromatography
- Dynamic light scattering (DLS)
- Small-angle X-ray scattering (SAXS)
Main Results:
- Determined the X-ray crystal structure of E. coli PmrD.
- Identified unambiguous secondary structures and a novel disulfide bond.
- Elucidated the structural and functional significance of the disulfide bond through various biochemical assays.
Conclusions:
- Structural characteristics of E. coli PmrD were clearly identified.
- The findings explain the unique polymyxin resistance mechanism in E. coli.
- Provides insights into differential resistance strategies among Gram-negative bacteria.
