Dataset Information

Structure-function of cyanobacterial outer-membrane protein, Slr1270: homolog of Escherichia coli drug export/colicin import protein, TolC.

ABSTRACT: Compared to thylakoid and inner membrane proteins in cyanobacteria, no structure-function information is available presently for integral outer-membrane proteins (OMPs). The Slr1270 protein from the cyanobacterium Synechocystis 6803, over-expressed in Escherichia coli, was refolded, and characterized for molecular size, secondary structure, and ion-channel function. Refolded Slr1270 displays a single band in native-electrophoresis, has an ?-helical content of 50-60%, as in E. coli TolC with which it has significant secondary-structure similarity, and an ion-channel function with a single-channel conductance of 80-200pS, and a monovalent ion (K(+):Cl(-)) selectivity of 4.7:1. The pH-dependence of channel conductance implies a role for carboxylate residues in channel gating, analogous to that in TolC.

SUBMITTER: Agarwal R

PROVIDER: S-EPMC4288923 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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Structure-function of cyanobacterial outer-membrane protein, Slr1270: homolog of Escherichia coli drug export/colicin import protein, TolC.

Agarwal Rachna R Zakharov Stanislav S Hasan S Saif SS Ryan Christopher M CM Whitelegge Julian P JP Cramer William A WA

FEBS letters 20140913 21

Compared to thylakoid and inner membrane proteins in cyanobacteria, no structure-function information is available presently for integral outer-membrane proteins (OMPs). The Slr1270 protein from the cyanobacterium Synechocystis 6803, over-expressed in Escherichia coli, was refolded, and characterized for molecular size, secondary structure, and ion-channel function. Refolded Slr1270 displays a single band in native-electrophoresis, has an α-helical content of 50-60%, as in E. coli TolC with whic ...[more]

PMID: 25218435

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Project description:Colicins are protein toxins made by Escherichia coli to kill related bacteria that compete for scarce resources. All colicins must cross the target cell outer membrane in order to reach their intracellular targets. Normally, the first step in the intoxication process is the tight binding of the colicin to an outer membrane receptor protein via its central receptor-binding domain. It is shown here that for one colicin, E1, that step, although it greatly increases the efficiency of killing, is not absolutely necessary. For colicin E1, the second step, translocation, relies on the outer membrane/transperiplasmic protein TolC. The normal role of TolC in bacteria is as an essential component of a family of tripartite drug and toxin exporters, but for colicin E1, it is essential for its import. Colicin E1 and some N-terminal translocation domain peptides had been shown previously to bind in vitro to TolC and occlude channels made by TolC in planar lipid bilayer membranes. Here, a set of increasingly shorter colicin E1 translocation domain peptides was shown to bind to Escherichia coli in vivo and protect them from subsequent challenge by colicin E1. A segment of only 21 residues, the "TolC box," was thereby defined; that segment is essential for colicin E1 cytotoxicity and for binding of translocation domain peptides to TolC. IMPORTANCE:The Escherichia coli outer membrane/transperiplasmic protein TolC is normally an essential component of the bacterium's tripartite drug and toxin export machinery. The protein toxin colicin E1 instead uses TolC for its import into the cells that it kills, thereby subverting its normal role. Increasingly shorter constructs of the colicin's N-terminal translocation domain were used to define an essential 21-residue segment that is required for both colicin cytotoxicity and for binding of the colicin's translocation domain to bacteria, in order to protect them from subsequent challenge by active colicin E1. Thus, an essential TolC binding sequence of colicin E1 was identified and may ultimately lead to the development of drugs to block the bacterial drug export pathway.

Dataset Information

Structure-function of cyanobacterial outer-membrane protein, Slr1270: homolog of Escherichia coli drug export/colicin import protein, TolC.

Publications

Structure-function of cyanobacterial outer-membrane protein, Slr1270: homolog of Escherichia coli drug export/colicin import protein, TolC.

Similar Datasets

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