TY - JOUR
T1 - Expression, purification and crystallization of a cohesin domain from the cellulosome of Clostridium thermocellum
AU - Yaron, Sima
AU - Shimon, Linda J.W.
AU - Frolow, Felix
AU - Lamed, Raphael
AU - Morag, Ely
AU - Shoham, Yuval
AU - Bayer, Edward A.
N1 - Funding Information:
The authors acknowledge the technical help of Dorit Nahari in purifying the recombinant co-hesin domain. This research was supported by the Israel Science Foundation, administered by the Israel Academy of Sciences and Humanities. Technical support was provided by the Technion Otto Meyerhof Biotechnology Laboratories established by the Minerva Foundation, Germany.
PY - 1996/11/15
Y1 - 1996/11/15
N2 - The cellulosome of the cellulolytic bacterium, Clostridium thermocellum, is a multi-enzyme complex in which the enzymatic (cellulolytic) subunits are attached to a unique nonhydrolytic subunit called scaffoldin. The attachment is mediated by two mutually interacting domains: namely, multiple cohesin domains on the scaffoldin subunit and a dockerin domain on each of the enzymatic subunits. Knowledge of the three-dimensional structure of each of the interacting components would be critical to a better understanding of the cohesin-dockerin interaction at the molecular level. In this report, we describe the purification of one of the nine cohesin domains of the scaffoldin subunit from C. thermocellum. A DNA segment containing the cohesin 2 sequence was fused to a hexa-histidine tag, and the resultant construct was expressed in Escherichia coli. The expressed peptide was efficiently isolated by metal-chelate affinity chromatography. The purified recombinant form of the cohesin was crystallized pending determination of its structure.
AB - The cellulosome of the cellulolytic bacterium, Clostridium thermocellum, is a multi-enzyme complex in which the enzymatic (cellulolytic) subunits are attached to a unique nonhydrolytic subunit called scaffoldin. The attachment is mediated by two mutually interacting domains: namely, multiple cohesin domains on the scaffoldin subunit and a dockerin domain on each of the enzymatic subunits. Knowledge of the three-dimensional structure of each of the interacting components would be critical to a better understanding of the cohesin-dockerin interaction at the molecular level. In this report, we describe the purification of one of the nine cohesin domains of the scaffoldin subunit from C. thermocellum. A DNA segment containing the cohesin 2 sequence was fused to a hexa-histidine tag, and the resultant construct was expressed in Escherichia coli. The expressed peptide was efficiently isolated by metal-chelate affinity chromatography. The purified recombinant form of the cohesin was crystallized pending determination of its structure.
KW - Clostridium thermocellum
KW - X-ray crystallography
KW - cellulases
KW - cellulosome
KW - cohesin domain
UR - http://www.scopus.com/inward/record.url?scp=0030588690&partnerID=8YFLogxK
U2 - 10.1016/S0168-1656(96)01602-1
DO - 10.1016/S0168-1656(96)01602-1
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AN - SCOPUS:0030588690
SN - 0168-1656
VL - 51
SP - 243
EP - 249
JO - Journal of Biotechnology
JF - Journal of Biotechnology
IS - 3
T2 - Proceedings of the 1995 Workshop on Environmental Biotechnology
Y2 - 28 November 1995 through 28 November 1995
ER -