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Journal of Bacteriology, March 1999, p. 1801-1810, Vol. 181, No. 6
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Sequence Analysis of Scaffolding Protein CipC and
ORFXp, a New Cohesin-Containing Protein in
Clostridium cellulolyticum: Comparison of Various
Cohesin Domains and Subcellular Localization of ORFXp
Sandrine
Pagès,1
Anne
Bélaïch,1,*
Henri-Pierre
Fierobe,1
Chantal
Tardif,1,2
Christian
Gaudin,1 and
Jean-Pierre
Bélaïch1,2
Bioénergétique et
Ingéniérie des Protéines, Centre National de la
Recherche Scientifique,1 and
Université de Provence,2
Marseilles, France
Received 8 September 1998/Accepted 6 January 1999
The gene encoding the scaffolding protein of the cellulosome from
Clostridium cellulolyticum, whose partial sequence was
published earlier (S. Pagès, A. Bélaïch, C. Tardif,
C. Reverbel-Leroy, C. Gaudin, and J.-P. Bélaïch, J. Bacteriol. 178:2279-2286, 1996; C. Reverbel-Leroy, A. Bélaïch, A. Bernadac, C. Gaudin, J. P. Bélaïch, and C. Tardif, Microbiology 142:1013-1023,
1996), was completely sequenced. The corresponding protein, CipC, is
composed of a cellulose binding domain at the N terminus followed by
one hydrophilic domain (HD1), seven highly homologous cohesin domains (cohesin domains 1 to 7), a second hydrophilic domain, and a final cohesin domain (cohesin domain 8) which is only 57 to 60% identical to
the seven other cohesin domains. In addition, a second gene located
8.89 kb downstream of cipC was found to encode a
three-domain protein, called ORFXp, which includes a cohesin domain. By
using antiserum raised against the latter, it was observed that ORFXp is associated with the membrane of C. cellulolyticum and is
not detected in the cellulosome fraction. Western blot and BIAcore experiments indicate that cohesin domains 1 and 8 from CipC recognize the same dockerins and have similar affinity for CelA
(Ka = 4.8 × 109
M
1) whereas the cohesin from ORFXp, although it is also
able to bind all cellulosome components containing a dockerin, has a
19-fold lower Ka for CelA (2.6 × 108 M
1). Taken together, these data suggest
that ORFXp may play a role in cellulosome assembly.
*
Corresponding author. Mailing address:
Bioénergétique et Ingéniérie des
Protéines, Centre National de la Recherche Scientifique, 31 chemin Joseph Aiguier, BP 71, 13402 Marseilles Cedex 20, France. Phone:
(33) 91 16 40 70. Fax: (33) 91 71 33 21. E-mail:
abelaich{at}ibsm.cnrs-mrs.fr.
Journal of Bacteriology, March 1999, p. 1801-1810, Vol. 181, No. 6
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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