Amino Acid Residues Involved in Membrane Insertion and Pore Formation of Clostridium botulinum C2 Toxin
Identifieur interne : 001952 ( Main/Exploration ); précédent : 001951; suivant : 001953Amino Acid Residues Involved in Membrane Insertion and Pore Formation of Clostridium botulinum C2 Toxin
Auteurs : Alexander E. Lang [États-Unis] ; Tobias Neumeyer [États-Unis] ; Jianjun Sun [États-Unis] ; R. John Collier [États-Unis] ; Roland Benz [États-Unis] ; Klaus Aktories [États-Unis, Allemagne]Source :
- Biochemistry [ 0006-2960 ] ; 2008.
Abstract
The actin-ADP-ribosylating Clostridium botulinum C2 toxin consists of the enzymatic component C2I and the binding component C2II. C2II forms heptameric channels involved in translocation of the enzymatic component into the target cell. On the basis of the heptameric toxin channel, we studied functional consequences of mutagenesis of amino acid residues probably lining the lumen of the toxin channel. Substitution of glutamate-399 of C2II with alanine blocked channel formation and cytotoxicity of the holotoxin. Although cytotoxicity and rounding up of cells by C2I were completely blocked by exchange of phenylalanine-428 with alanine, the mutation increased potassium conductance caused by C2II in artificial membranes by about 2−3-fold over that of wild-type toxin. In contrast to its effects on single-channel potassium conductance in artificial membranes, the F428A mutation delayed the kinetics of pore formation in lipid vesicles and inhibited the activity of C2II in promoting 86Rb+ release from preloaded intact cells after pH shift of the medium. Moreover, F428A C2II exhibited delayed and diminished formation of C2II aggregates at low pH, indicating major changes of the biophysical properties of the toxin. The data indicate that phenylalanine-428 of C2II plays a major role in conformational changes occurring during pore formation of the binding component of C2II.
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DOI: 10.1021/bi800615g
Affiliations:
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John Collier</name></author><author><name sortKey="Benz, Roland" sort="Benz, Roland" uniqKey="Benz R" first="Roland" last="Benz">Roland Benz</name></author><author><name sortKey="Aktories, Klaus" sort="Aktories, Klaus" uniqKey="Aktories K" first="Klaus" last="Aktories">Klaus Aktories</name></author></titleStmt><publicationStmt><idno type="wicri:source">ISTEX</idno><idno type="RBID">ISTEX:EB9BE043BD5AB987E9AF8EE5527CF2291E23487D</idno><date when="2008" year="2008">2008</date><idno type="doi">10.1021/bi800615g</idno><idno type="url">https://api.istex.fr/ark:/67375/TPS-NMLWNX7Z-G/fulltext.pdf</idno><idno type="wicri:Area/Istex/Corpus">002005</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Corpus" wicri:corpus="ISTEX">002005</idno><idno type="wicri:Area/Istex/Curation">002005</idno><idno type="wicri:Area/Istex/Checkpoint">000857</idno><idno type="wicri:explorRef" wicri:stream="Istex" wicri:step="Checkpoint">000857</idno><idno type="wicri:doubleKey">0006-2960:2008:Lang A:amino:acid:residues</idno><idno type="wicri:Area/Main/Merge">001960</idno><idno type="wicri:Area/Main/Curation">001952</idno><idno type="wicri:Area/Main/Exploration">001952</idno></publicationStmt><sourceDesc><biblStruct><analytic><title level="a" type="main" xml:lang="en">Amino Acid Residues Involved in Membrane Insertion and Pore Formation of <hi rend="italic">Clostridium botulinum</hi> C2 Toxin<ref type="bib" target="#fn1"></ref></title><author><name sortKey="Lang, Alexander E" sort="Lang, Alexander E" uniqKey="Lang A" first="Alexander E." last="Lang">Alexander E. Lang</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation></author><author><name sortKey="Neumeyer, Tobias" sort="Neumeyer, Tobias" uniqKey="Neumeyer T" first="Tobias" last="Neumeyer">Tobias Neumeyer</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation></author><author><name sortKey="Sun, Jianjun" sort="Sun, Jianjun" uniqKey="Sun J" first="Jianjun" last="Sun">Jianjun Sun</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation></author><author><name sortKey="Collier, R John" sort="Collier, R John" uniqKey="Collier R" first="R. John" last="Collier">R. John Collier</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation></author><author><name sortKey="Benz, Roland" sort="Benz, Roland" uniqKey="Benz R" first="Roland" last="Benz">Roland Benz</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation></author><author><name sortKey="Aktories, Klaus" sort="Aktories, Klaus" uniqKey="Aktories K" first="Klaus" last="Aktories">Klaus Aktories</name><affiliation wicri:level="2"><country xml:lang="fr">États-Unis</country><placeName><region type="state">Massachusetts</region></placeName><wicri:cityArea>Institut für Experimentelle and Klinische Pharmakologie and Toxikologie, Albert-Ludwigs-Universität Freiburg, Albertstrasse 25, D-79104 Freiburg, Germany, Lehrstuhl für Biotechnologie, Theodor-Boveri-Institut (Biozentrum), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany, and Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, Boston</wicri:cityArea></affiliation><affiliation></affiliation><affiliation wicri:level="1"><country wicri:rule="url">Allemagne</country></affiliation></author></analytic><monogr></monogr><series><title level="j" type="main">Biochemistry</title><title level="j" type="abbrev">Biochemistry</title><idno type="ISSN">0006-2960</idno><idno type="eISSN">1520-4995</idno><imprint><publisher>American Chemical Society</publisher><date type="e-published" when="2008-07-18">2008</date><date when="2008-08-12">2008</date><biblScope unit="vol">47</biblScope><biblScope unit="issue">32</biblScope><biblScope unit="page" from="8406">8406</biblScope><biblScope unit="page" to="8413">8413</biblScope></imprint><idno type="ISSN">0006-2960</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0006-2960</idno></seriesStmt></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract">The actin-ADP-ribosylating Clostridium botulinum C2 toxin consists of the enzymatic component C2I and the binding component C2II. C2II forms heptameric channels involved in translocation of the enzymatic component into the target cell. On the basis of the heptameric toxin channel, we studied functional consequences of mutagenesis of amino acid residues probably lining the lumen of the toxin channel. Substitution of glutamate-399 of C2II with alanine blocked channel formation and cytotoxicity of the holotoxin. Although cytotoxicity and rounding up of cells by C2I were completely blocked by exchange of phenylalanine-428 with alanine, the mutation increased potassium conductance caused by C2II in artificial membranes by about 2−3-fold over that of wild-type toxin. In contrast to its effects on single-channel potassium conductance in artificial membranes, the F428A mutation delayed the kinetics of pore formation in lipid vesicles and inhibited the activity of C2II in promoting 86Rb+ release from preloaded intact cells after pH shift of the medium. Moreover, F428A C2II exhibited delayed and diminished formation of C2II aggregates at low pH, indicating major changes of the biophysical properties of the toxin. The data indicate that phenylalanine-428 of C2II plays a major role in conformational changes occurring during pore formation of the binding component of C2II.</div></front></TEI><affiliations><list><country><li>Allemagne</li><li>États-Unis</li></country><region><li>Massachusetts</li></region></list><tree><country name="États-Unis"><region name="Massachusetts"><name sortKey="Lang, Alexander E" sort="Lang, Alexander E" uniqKey="Lang A" first="Alexander E." last="Lang">Alexander E. Lang</name></region><name sortKey="Aktories, Klaus" sort="Aktories, Klaus" uniqKey="Aktories K" first="Klaus" last="Aktories">Klaus Aktories</name><name sortKey="Benz, Roland" sort="Benz, Roland" uniqKey="Benz R" first="Roland" last="Benz">Roland Benz</name><name sortKey="Collier, R John" sort="Collier, R John" uniqKey="Collier R" first="R. John" last="Collier">R. John Collier</name><name sortKey="Neumeyer, Tobias" sort="Neumeyer, Tobias" uniqKey="Neumeyer T" first="Tobias" last="Neumeyer">Tobias Neumeyer</name><name sortKey="Sun, Jianjun" sort="Sun, Jianjun" uniqKey="Sun J" first="Jianjun" last="Sun">Jianjun Sun</name></country><country name="Allemagne"><noRegion><name sortKey="Aktories, Klaus" sort="Aktories, Klaus" uniqKey="Aktories K" first="Klaus" last="Aktories">Klaus Aktories</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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