The location of the ligand-binding site of carbohydrate-binding modules that have evolved from a common sequence is not conserved.
Polysaccharide-degrading enzymes are generally modular proteins that contain non-catalytic carbohydrate-binding modules (CBMs), which potentiate the activity of the catalytic module. CBMs have been grouped into sequence-based families, and three-dimensional structural data are available for half of these families. Clostridium thermocellum xylanase 11A is a modular enzyme that contains a CBM from family 6 (CBM6), for which no structural data are available. We have determined the crystal structure of this module to a resolution of 2.1 A. The protein is a beta-sandwich that contains two potential ligand-binding clefts designated cleft A and B. The CBM interacts primarily with xylan, and NMR spectroscopy coupled with site-directed mutagenesis identified cleft A, containing Trp-92, Tyr-34, and Asn-120, as the ligand-binding site. The overall fold of CBM6 is similar to proteins in CBM families 4 and 22, although surprisingly the ligand-binding site in CBM4 and CBM22 is equivalent to cleft B in CBM6. These structural data define a superfamily of CBMs, comprising CBM4, CBM6, and CBM22, and demonstrate that, although CBMs have evolved from a relatively small number of ancestors, the structural elements involved in ligand recognition have been assembled at different locations on the ancestral scaffold.
Czjzek
M
M
Laboratoire d'Architecture et de Fonction des Macromolécules Biologiques, IBSM, CNRS Marseille and University Aix-Marseille I & II, 31 Chemin Joseph Aiguier, 13402 Marseille Cedex 20, France. czjzek@afmb.cnrs-mrs.fr
Bolam
D N
DN
Mosbah
A
A
Allouch
J
J
Fontes
C M
CM
Ferreira
L M
LM
Bornet
O
O
Zamboni
V
V
Darbon
H
H
Smith
N L
NL
Black
G W
GW
Henrissat
B
B
Gilbert
H J
HJ
D000595:Amino Acid Sequence; D001665:Binding Sites; D050260:Carbohydrate Metabolism; D018360:Crystallography, X-Ray; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D043325:Xylan Endo-1,3-beta-Xylosidase; D014995:Xylosidases / Q000737:chemistry / Q000378:metabolism*
D008024:Ligands; D014995:Xylosidases; D043325:Xylan Endo-1,3-beta-Xylosidase