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Crystal structure of a COG4313 outer membrane channel.

van den Berg B, Bhamidimarri SP, Winterhalter M - Sci Rep (2015)

Bottom Line: Structure-function studies of this protein family have so far been hampered by a lack of structural information.Instead, the presence of bound detergent molecules inside the barrel suggests that Pput2725 mediates uptake of hydrophobic molecules.Sequence alignments and the locations of highly conserved residues suggest the presence of a dynamic lateral opening through which hydrophobic molecules might gain entry into the cell.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell and Molecular Biosciences, The Medical School, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK.

ABSTRACT
COG4313 proteins form a large and widespread family of outer membrane channels and have been implicated in the uptake of a variety of hydrophobic molecules. Structure-function studies of this protein family have so far been hampered by a lack of structural information. Here we present the X-ray crystal structure of Pput2725 from the biodegrader Pseudomonas putida F1, a COG4313 channel of unknown function, using data to 2.3 Å resolution. The structure shows a 12-stranded barrel with an N-terminal segment preceding the first β-strand occluding the lumen of the barrel. Single channel electrophysiology and liposome swelling experiments suggest that while the narrow channel visible in the crystal structure does allow passage of ions and certain small molecules in vitro, Pput2725 is unlikely to function as a channel for hydrophilic molecules. Instead, the presence of bound detergent molecules inside the barrel suggests that Pput2725 mediates uptake of hydrophobic molecules. Sequence alignments and the locations of highly conserved residues suggest the presence of a dynamic lateral opening through which hydrophobic molecules might gain entry into the cell. Our results provide the basis for structure-function studies of COG4313 family members with known function, such as the SphA sphingosine uptake channel of Pseudomonas aeruginosa.

No MeSH data available.


Related in: MedlinePlus

Detergent binding by Pput2725.Cartoon overviews of Pput2725 from the side and from the outside of the cell, showing the two C8E4 detergent molecules (det1 and det2) bound in the structure (carbons, cyan; oxygens, red). The N-terminus is colored green and loop L3 is shown in orange.
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f4: Detergent binding by Pput2725.Cartoon overviews of Pput2725 from the side and from the outside of the cell, showing the two C8E4 detergent molecules (det1 and det2) bound in the structure (carbons, cyan; oxygens, red). The N-terminus is colored green and loop L3 is shown in orange.

Mentions: What then is the transport substrate for Pput2725? Since the function of this protein is not yet known, it is not possible to answer this question conclusively. However, the crystal structure does provide a clue as to the nature of the substrate. Two tubular stretches of density are present in the Pput2725 structure, and these likely correspond to partially ordered C8E4 detergent molecules used for crystallisation (Fig. 4). One of the detergent molecules (det1) is bound in the external vestibule, with the other (det2) bound on the periplasmic side below the constriction. Our results therefore suggest that the transport substrate of Pput2725 is amphipathic, analogous to that of TcpY and SphA.


Crystal structure of a COG4313 outer membrane channel.

van den Berg B, Bhamidimarri SP, Winterhalter M - Sci Rep (2015)

Detergent binding by Pput2725.Cartoon overviews of Pput2725 from the side and from the outside of the cell, showing the two C8E4 detergent molecules (det1 and det2) bound in the structure (carbons, cyan; oxygens, red). The N-terminus is colored green and loop L3 is shown in orange.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493636&req=5

f4: Detergent binding by Pput2725.Cartoon overviews of Pput2725 from the side and from the outside of the cell, showing the two C8E4 detergent molecules (det1 and det2) bound in the structure (carbons, cyan; oxygens, red). The N-terminus is colored green and loop L3 is shown in orange.
Mentions: What then is the transport substrate for Pput2725? Since the function of this protein is not yet known, it is not possible to answer this question conclusively. However, the crystal structure does provide a clue as to the nature of the substrate. Two tubular stretches of density are present in the Pput2725 structure, and these likely correspond to partially ordered C8E4 detergent molecules used for crystallisation (Fig. 4). One of the detergent molecules (det1) is bound in the external vestibule, with the other (det2) bound on the periplasmic side below the constriction. Our results therefore suggest that the transport substrate of Pput2725 is amphipathic, analogous to that of TcpY and SphA.

Bottom Line: Structure-function studies of this protein family have so far been hampered by a lack of structural information.Instead, the presence of bound detergent molecules inside the barrel suggests that Pput2725 mediates uptake of hydrophobic molecules.Sequence alignments and the locations of highly conserved residues suggest the presence of a dynamic lateral opening through which hydrophobic molecules might gain entry into the cell.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell and Molecular Biosciences, The Medical School, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK.

ABSTRACT
COG4313 proteins form a large and widespread family of outer membrane channels and have been implicated in the uptake of a variety of hydrophobic molecules. Structure-function studies of this protein family have so far been hampered by a lack of structural information. Here we present the X-ray crystal structure of Pput2725 from the biodegrader Pseudomonas putida F1, a COG4313 channel of unknown function, using data to 2.3 Å resolution. The structure shows a 12-stranded barrel with an N-terminal segment preceding the first β-strand occluding the lumen of the barrel. Single channel electrophysiology and liposome swelling experiments suggest that while the narrow channel visible in the crystal structure does allow passage of ions and certain small molecules in vitro, Pput2725 is unlikely to function as a channel for hydrophilic molecules. Instead, the presence of bound detergent molecules inside the barrel suggests that Pput2725 mediates uptake of hydrophobic molecules. Sequence alignments and the locations of highly conserved residues suggest the presence of a dynamic lateral opening through which hydrophobic molecules might gain entry into the cell. Our results provide the basis for structure-function studies of COG4313 family members with known function, such as the SphA sphingosine uptake channel of Pseudomonas aeruginosa.

No MeSH data available.


Related in: MedlinePlus