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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

The location of conserved residues in Pput2725.(A) Stereo diagram viewed from the side, showing identical residues as magenta stick models (nitrogen atoms; blue, oxygen atoms; red). The N-terminus is colored green. (B) Surface model in a slightly different orientation relative to (A). Aromatic residues delineating the approximate external hydrophobic-hydrophilic interface of the OM (dashed line) are colored yellow.
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f6: The location of conserved residues in Pput2725.(A) Stereo diagram viewed from the side, showing identical residues as magenta stick models (nitrogen atoms; blue, oxygen atoms; red). The N-terminus is colored green. (B) Surface model in a slightly different orientation relative to (A). Aromatic residues delineating the approximate external hydrophobic-hydrophilic interface of the OM (dashed line) are colored yellow.

Mentions: When the remaining fifteen identical residues are highlighted onto the Pput2725 crystal structure, a remarkable pattern emerges. No fewer than nine residues in loops L2, L3 and L4 are clustered together (Asp92, Thr94, Pro119, Thr120, Gly121, Tyr123, Asn130, Asn134 and Asn166) (Fig. 6), hinting at a functional importance of this region. Most of the polar side chains appear to be involved in interactions stabilising the non-β-strand conformations of the loops. Viewed on a molecular surface model the conserved residues are clustered together and form a striking circular patch. The presence of the conserved cluster may have important implications for the possible transport mechanism, which will be discussed below.


Crystal structure of a COG4313 outer membrane channel.

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

The location of conserved residues in Pput2725.(A) Stereo diagram viewed from the side, showing identical residues as magenta stick models (nitrogen atoms; blue, oxygen atoms; red). The N-terminus is colored green. (B) Surface model in a slightly different orientation relative to (A). Aromatic residues delineating the approximate external hydrophobic-hydrophilic interface of the OM (dashed line) are colored yellow.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f6: The location of conserved residues in Pput2725.(A) Stereo diagram viewed from the side, showing identical residues as magenta stick models (nitrogen atoms; blue, oxygen atoms; red). The N-terminus is colored green. (B) Surface model in a slightly different orientation relative to (A). Aromatic residues delineating the approximate external hydrophobic-hydrophilic interface of the OM (dashed line) are colored yellow.
Mentions: When the remaining fifteen identical residues are highlighted onto the Pput2725 crystal structure, a remarkable pattern emerges. No fewer than nine residues in loops L2, L3 and L4 are clustered together (Asp92, Thr94, Pro119, Thr120, Gly121, Tyr123, Asn130, Asn134 and Asn166) (Fig. 6), hinting at a functional importance of this region. Most of the polar side chains appear to be involved in interactions stabilising the non-β-strand conformations of the loops. Viewed on a molecular surface model the conserved residues are clustered together and form a striking circular patch. The presence of the conserved cluster may have important implications for the possible transport mechanism, which will be discussed below.

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