Gating processes of channels induced by colicin A, its C-terminal fragment and colicin E1 in planar lipid bilayers

Collarini, M; Amblard, G.; Lazdunski, Claude; Pattus, Franc

doi:10.1007/bf00253839

Cited by 64 publications

(53 citation statements)

References 27 publications

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“…Analogous to other systems [19,21], these increments can be attributed to incorporation and activation of pores formed by maltoporin within the bilayer. After induction, the current reaches a steady state where fluctuations occur frequently ( fig.…”

Section: Maltoporin Channel Conductancementioning

confidence: 81%

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Selectivity for maltose and maltodextrins of maltoporin, a pore‐forming protein of E. coli outer membrane

Dargent¹,

Rosenbusch

Pattus³

1987

FEBS Letters

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Homogeneous maltoporin (1amB protein), an Escherichia coli outer membrane spanning protein, was incorporated in phospholipid planar bilayers. It generates aqueous channels distinct from those formed by the non-specific porin (OmpF) or by phosphoporin (phoE protein). The single conductance, 150 pS in 1 M NaCI, is much smaller than that of the porins. The channels, which are poorly selective for cations and voltage independent, are specifically inhibited by maltose and maltodextrins. This inhibition, observed in the absence of maltose binding protein, demonstrates that the selectivity of maltoporin for maltose and maltodextrins is an intrinsic property of the protein.

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Section: Maltoporin Channel Conductancementioning

confidence: 81%

“…Benz et al [13] Schindler [IS]; or (ii) highly purified maltoporin in detergent was injected in front of a preformed bilayer obtained as above as described by Pattus and co-workers [19]. By using this last experimental approach, the number of induced channels varies 2-or 3-fold.…”

Section: Introductionmentioning

confidence: 99%

Selectivity for maltose and maltodextrins of maltoporin, a pore‐forming protein of E. coli outer membrane

Dargent¹,

Rosenbusch

Pattus³

1987

FEBS Letters

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“…The decrease in conformational entropy when two protein domains are tightly coupled to each other is expected to contribute to the destabilization of large protein domains, but more specific interactions between the proteins, such as van der Waals, hydrogen bonding, or electrostatic interactions, can also be important. In a related manner, functional differences in ColA PFF and colicin A have been attributed to interactions between the pore-forming and the receptor binding domains (32,42).…”

Section: Structural and Functional Characteristics Of Colicin B Andmentioning

confidence: 99%

Calorimetric Investigations of the Structural Stability and Interactions of Colicin B Domains in Aqueous Solution and in the Presence of Phospholipid Bilayers

Ortega¹,

Lambotte²,

Bechinger³

2001

Journal of Biological Chemistry

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The effects of pH and temperature on the stability of interdomain interactions of colicin B have been studied by differential-scanning calorimetry, circular dichroism, and fluorescence spectroscopy. The calorimetric properties were compared with those of the isolated pore-forming fragment. The unfolding profile of the fulllength toxin is consistent with two endothermic transitions. Whereas peak A (T m ‫؍‬ 55°C) most likely corresponds to the receptor/translocation domain, peak B (T m ‫؍‬ 59°C) is associated with the pore-forming domain. By lowering the pH from 7 to 3.5, the transition temperature of peaks A and B are reduced by 25 and 18°C, respectively, due to proton exchange upon denaturation. The isolated pore-forming fragment unfolds at much higher temperatures (T m ‫؍‬ 65°C) and is stable throughout a wide pH range, indicating that intramolecular interactions between the different colicin B domains result in a less stable protein conformation. In aqueous solution circular dichroism spectra have been used to estimate the content of helical secondary structure of colicin B (Ϸ40%) or its pore-forming fragment (Ϸ80%). Upon heating, the ellipticities at 222 nm strongly decrease at the transition temperature. In the presence of lipid vesicles the differential-scanning calorimetry profiles of the pore-forming fragment exhibit a low heat of transition multicomponent structure. The heat of transition of membrane-associated colicin B (T m ‫؍‬ 54°C at pH 3.5) is reduced and its secondary structure is conserved even at intermediate temperatures indicating incomplete unfolding due to strong protein-lipid interactions.

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“…In order to study this voltage dependence on a sample of many channels, quasi-steady-state conditions were achieved by holding the membrane at f90 mV for 1 h until the rate of new channel incorporation was very low. In the past, two measurements have been used to define the voltage dependence of colicin A and of its C-terminal fragment [19]: the steady state, macroscopic, current versus voltage curve and the voltage dependence of the time constants of relaxation following a voltage step. The voltage steps used in this study were from + 100 mV, at which point all channels were open, to a less positive value of VH.…”

Section: The Da70c Fragment Can Form Voltage-dependent Channelsmentioning

confidence: 99%

A 136‐amino‐acid‐residue COOH‐terminal fragment of colicin A is endowed with ionophoric activity

Baty¹,

Lakey

Pattus

et al. 1990

European Journal of Biochemistry

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DNA regions encoding the various domains of a protein can be expressed as separate entities by inserting at appropriate sites a 'STOP-Shine-Dalgarno-sequence-ATG' cassette encoding a termination codon, a ShineDalgano sequence and an initiation codon within the structural gene. This technique has been used to obtain a 137-amino-acid-residue pore-forming protein designated DA70C comprising the final 136-amino-acid-residue COOH-terminal of colicin A preceded by an NH,-terminal methionine. Da70C was correctly expressed but poorly released to the extracellular medium. Its purification involved, as a final step, a partition in Triton X-114 thus demonstrating that hydrophobic regions are exposed in this protein. The ability of DA70C to form ion channels in planar lipid bilayers was investigated and pore properties were analyzed. The results indicate that helices 1 -3 of the 204-amino-acid-residue colicin pore-forming domain (containing 10 a-heIices) are not involved in ion conduction through the channel. However, they are important in maintaining the stability of the soluble state of the COOH-terminal domain.Colicin A is a bacterial toxin produced by Escherichia coli and closely related bacteria. It belongs to the largest group of colicins comprising those which can form voltage-dependent channels in membranes, thereby destroying the cell's energy potential [l].The channel forming domain is contained in the carboxy-terminal region of the molecule [2]. A 204-aminoacid-residue COOH-terminal fragment obtained by digestion with thermolysin comprising amino acids 389 -592 from colicin A has pore-forming properties similar to those of the entire molecule [3]. This fragment has the unusual property to be both soluble in aqueous medium and able spontaneously to insert into lipid monolayers and bilayers [4]. The threedimensional structure at 25-fm resolution has been recently determined [4]. The protein consists of ten a-helices organized in a three-layer structure. Two of these helices are entirely hydrophilic and completely buried within the structure of the 204-residue COOH-terminal fragment. The colicin El COOHterminal domain is significantly similar to colicin A [1,5 -71. The minimum size of this domain that is competent for channel function is under debate [8]. However, it is clear that a colicin El COOH-terminal fragment of 152 residues is competent for channel function [9]. This would mean that helices 1 and 2, recently identified in the three-dimensional structure of colicin A [4], are not required for channel formation. It has been reported that the a-helical content of the lipid-associated COOH-terminal fragment of colicin A is similar to that of the soluble form [5]. This suggests that about the same helices as that identified in the soluble form [4] exist in the membraneinserted form of this fragment.We have previously demonstrated that DNA regions encoding the various domains of a protein can be expressed as separate entities by inserting at appropriate sites a 'cassette' encoding a termination codon, a Shine-Dalg...

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Gating processes of channels induced by colicin A, its C-terminal fragment and colicin E1 in planar lipid bilayers

Cited by 64 publications

References 27 publications

Selectivity for maltose and maltodextrins of maltoporin, a pore‐forming protein of E. coli outer membrane

Selectivity for maltose and maltodextrins of maltoporin, a pore‐forming protein of E. coli outer membrane

Calorimetric Investigations of the Structural Stability and Interactions of Colicin B Domains in Aqueous Solution and in the Presence of Phospholipid Bilayers

A 136‐amino‐acid‐residue COOH‐terminal fragment of colicin A is endowed with ionophoric activity

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