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Journal of Bacteriology, May 2000, p. 2761-2770, Vol. 182, No. 10
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Enzymology of Type IV Macromolecule Secretion
Systems: the Conjugative Transfer Regions of Plasmids RP4 and R388 and
the cag Pathogenicity Island of Helicobacter
pylori Encode Structurally and Functionally Related Nucleoside
Triphosphate Hydrolases
Sabine
Krause,1
Werner
Pansegrau,2,
Rudi
Lurz,1
Fernando
de la
Cruz,3 and
Erich
Lanka1,*
Max-Planck-Institut für Molekulare
Genetik, D-14195 Berlin, Germany1;
Institute for Molecular Plant Sciences, Clusius
Laboratory, Leiden University, 2333 AL Leiden, The
Netherlands2; and Departamento de Biologia
Molecular, Facultad de Medicina, Universidad de Cantabria, s/n
39011 Santander, Spain3
Received 13 December 1999/Accepted 1 March 2000
Type IV secretion systems direct transport of protein or
nucleoprotein complexes across the cell envelopes of prokaryotic donor
and eukaryotic or prokaryotic recipient cells. The process is mediated
by a membrane-spanning multiprotein assembly. Potential NTPases
belonging to the VirB11 family are an essential part of the
membrane-spanning complex. Three representatives of these NTPases
originating from the conjugative transfer regions of plasmids RP4
(TrbB) and R388 (TrwD) and from the cag pathogenicity
island of Helicobacter pylori (HP0525) were overproduced
and purified in native form. The proteins display NTPase activity with
distinct substrate specificities in vitro. TrbB shows its highest
specific hydrolase activity with dATP, and the preferred substrate for HP0525 is ATP. Analysis of defined TrbB mutations altered in motifs conserved within the VirB11 protein family shows that there is a
correlation between the loss or reduction of NTPase activity and
transfer frequency. Tryptophan fluorescence spectroscopy of TrbB and
HP0525 suggests that both interact with phospholipid membranes,
changing their conformation. NTPase activity of both proteins was
stimulated by the addition of certain phospholipids. According to our
results, Virb11-like proteins seem to most likely be involved in the
assembly of the membrane-spanning multiprotein complex.
*
Corresponding author. Mailing address:
Max-Planck-Institut für Molekulare Genetik, Abteilung Lehrach,
Ihnestrasse 73, D-14195 Berlin, Germany. Phone: 49 30 8413 1696. Fax:
49 30 8413 1130. E-mail: lanka{at}molgen.mpg.de.

Present address: IRIS Research Center, Chiron S.p.A.,
I-53100 Siena,
Italy.
Journal of Bacteriology, May 2000, p. 2761-2770, Vol. 182, No. 10
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
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