The C-terminal half of the colicin A pore-forming domain is active in vivo and in vitro11Edited by I. B. Holland

Nardi, Angèle; Slatin, Stephen L.; Baty, Daniel; Duché, Denis

doi:10.1006/jmbi.2001.4524

Cited by 21 publications

(16 citation statements)

References 31 publications

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“…In the case of colicin Ia, the presence of H1 as a transmembrane element is an important determinant of channel properties (6,24), but in the present case it appears that channels prevented from translocating helices 1-3 are not critically altered. This result is, however, consistent with the observation that helices 1-4 are not important for the colicin A channel (13), and thus may reflect a difference between colicins A and Ia.…”

Section: Discussionsupporting

confidence: 90%

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Translocation of a functional protein by a voltage-dependent ion channel

Slatin

Nardi

Jakes

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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The voltage-dependent gating of the colicin channel involves a substantial structural rearrangement that results in the transfer of about 35% of the 200 residues in its pore-forming domain across the membrane. This transfer appears to represent an unusual type of protein translocation that does not depend on a large, multimeric, protein pore. To investigate the ability of this system to transport arbitrary proteins, we made use of a pair of strongly interacting proteins, either of which could serve as a translocated cargo or as a probe to detect the other. Here we show that both an 86-residue and a 134-residue hydrophilic protein inserted into the translocated segment of colicin A are themselves translocated and are functional on the trans side of the bilayer. The disparate features of these proteins suggest that the colicin channel has a general protein translocation mechanism.

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Section: Discussionsupporting

confidence: 90%

“…In colicin Ia, a region corresponding to helices 2, 3, 4, and 5 of the soluble structure crosses the bilayer in association with gating (6). Colicin A, used here, is thought to support a similar translocation function, although the exact extent of the translocated domain may differ somewhat from colicin Ia (13).…”

mentioning

confidence: 92%

Translocation of a functional protein by a voltage-dependent ion channel

Slatin

Nardi

Jakes

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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“…For both colicins E1 and A, several helices worth of protein can be completely eliminated from the N terminus of the domain without preventing channel formation (16,124,408,478). There remains some uncertainty about the precise location of the upstream edge of the channel, part of which results from the fact that some of the shorter fragments tested form channels with altered properties compared to the native protein and part of which may reflect true differences among the colicins.…”

Section: Pore-forming Colicinsmentioning

confidence: 99%

Colicin Biology

Cascales

Buchanan

Duché

et al. 2007

Microbiol Mol Biol Rev

985

1,324

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SUMMARY Colicins are proteins produced by and toxic for some strains of Escherichia coli. They are produced by strains of E. coli carrying a colicinogenic plasmid that bears the genetic determinants for colicin synthesis, immunity, and release. Insights gained into each fundamental aspect of their biology are presented: their synthesis, which is under SOS regulation; their release into the extracellular medium, which involves the colicin lysis protein; and their uptake mechanisms and modes of action. Colicins are organized into three domains, each one involved in a different step of the process of killing sensitive bacteria. The structures of some colicins are known at the atomic level and are discussed. Colicins exert their lethal action by first binding to specific receptors, which are outer membrane proteins used for the entry of specific nutrients. They are then translocated through the outer membrane and transit through the periplasm by either the Tol or the TonB system. The components of each system are known, and their implication in the functioning of the system is described. Colicins then reach their lethal target and act either by forming a voltage-dependent channel into the inner membrane or by using their endonuclease activity on DNA, rRNA, or tRNA. The mechanisms of inhibition by specific and cognate immunity proteins are presented. Finally, the use of colicins as laboratory or biotechnological tools and their mode of evolution are discussed.

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“…pVL1 encodes the wild-type Cai with the 178 first amino acids of ColA fused to its N terminus (20). pCT1 encodes the pore-forming domain of ColA fused to the PelB sequence signal (sp-pfColA) (21). pCT3 encodes the B3 mutant form of sp-pfColA (sp-pfColA B3 ) (21).…”

Section: Methodsmentioning

confidence: 99%

“…pCT1 encodes the pore-forming domain of ColA fused to the PelB sequence signal (sp-pfColA) (21). pCT3 encodes the B3 mutant form of sp-pfColA (sp-pfColA B3 ) (21). pBAD/HisC contains the araBAD promoter from E. coli and was used to insert the gene encoding sp-pfColA or sppfColAB3.…”

Section: Methodsmentioning

confidence: 99%

Channel Domain of Colicin A Modifies the Dimeric Organization of Its Immunity Protein

Zhang

Lloubès

Duché

2010

Journal of Biological Chemistry

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Proteins conferring immunity against pore-forming colicins are localized in the Escherichia coli inner membrane. Their protective effects are mediated by direct interaction with the C-terminal domain of their cognate colicins. Cai, the immunity protein protecting E. coli against colicin A, contains four cysteine residues. We report cysteine cross-linking experiments showing that Cai forms homodimers. Cai contains four transmembrane segments (TMSs), and dimerization occurs via the third TMS. Furthermore, we observe the formation of intramolecular disulfide bonds that connect TMS2 with either TMS1 or TMS3. Co-expression of Cai with its target, the colicin A pore-forming domain (pfColA), in the inner membrane prevents the formation of intermolecular and intramolecular disulfide bonds, indicating that pfColA interacts with the dimer of Cai and modifies its conformation. Finally, we show that when Cai is locked by disulfide bonds, it is no longer able to protect cells against exogenous added colicin A.Pore-forming colicins are plasmid-encoded bacteriocins, synthesized by Enterobacteriacae, that are lethal to other related strains. Like many toxins, colicins are organized into structural domains that perform different functions: the Nterminal and the central domains are involved in the transport through the Escherichia coli envelope, and the C-terminal domain carries the toxic activity of the protein (1).The three-dimensional structures of the soluble form of the pore-forming domains of colicins A, E1, Ia, B, and N share the same general architecture: a bundle of eight amphipathic ␣-helices surrounding two hydrophobic ␣-helices (H8ϩH9), completely buried within the protein (2-6). However, crystal structures do not reveal the structure of the channel in the membrane, which remains uncharacterized. The hydrophobic helical hairpin may be the primary attachment region for channel formation and channel opening and closing require insertion and extrusion of an amphipathic segment, which has not been precisely defined (7,8).Each colicin plasmid also encodes a specific immunity protein that protects the producing cell against the cytotoxic activity of its colicin. Immunity proteins are integral inner membrane proteins. They are classified into two groups according to sequence similarities: the A type (immunity proteins to colicins A, B, N, and U) and the E1 type (immunity proteins to colicins E1, 5, K, 10, Ia, and Ib) (9 -11). The colicin A immunity protein (Cai) 2 has four transmembrane segments (TMSs), and its N and C termini are in the cytoplasm (Fig. 1), whereas the immunity protein to colicin E1 (Cei) crosses the cytoplasmic membrane three times, with the N terminus located in the cytoplasm and the C terminus in the periplasm (10, 11).The immunity proteins to pore-forming colicins diffuse laterally in the membrane and interact with helices of the pore-forming domain of their cognate colicin just prior channel opening (12-15). Genetic studies of A-type colicins indicate that the main determinant recognized by the immuni...

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The C-terminal half of the colicin A pore-forming domain is active in vivo and in vitro11Edited by I. B. Holland

Cited by 21 publications

References 31 publications

Translocation of a functional protein by a voltage-dependent ion channel

Translocation of a functional protein by a voltage-dependent ion channel

Colicin Biology

Channel Domain of Colicin A Modifies the Dimeric Organization of Its Immunity Protein

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