Recognition and hydrolysis of noncrystalline cellulose

被引:86
作者
Boraston, AB
Kwan, E
Chiu, P
Warren, RAJ
Kilburn, DG
机构
[1] PENCE Inc, Prot Engn Network Ctr Excellenc, Natl Business Ctr, Edmonton, AB T6G 2S2, Canada
[2] Univ British Columbia, Dept Microbiol & Immunol, Vancouver, BC V6T 1Z3, Canada
[3] Univ British Columbia, Biotechnol Lab, Vancouver, BC V6T 1Z3, Canada
关键词
D O I
10.1074/jbc.M209554200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Cellulase Ce15A from alkalophilic Bacillus sp. 1139 contains a family 17 carbohydrate-binding module (BspCBM17) and a family 28 CBM (BspCBM28) in tandem. The two modules have significantly similar amino acid sequences, but amino acid residues essential for binding are not conserved. BspCBM28 was obtained as a discrete polypeptide by engineering the cel5A gene. BspCBM17 could not be obtained as a discrete polypeptide, so a family 17 CBM from endoglucanase Cel5A of Clostridium cellulovorans, CcCBM17, was used to compare the binding characteristics of the two families of CBM. Both CcCBM17 and BspCBM28 recognized two classes of binding sites on amorphous cellulose: a high affinity site (K-a similar to1 x 10(6) m(-1)) and a low affinity site ( similar to2 X 10(4) m(-1)). They did not compete for binding to the high affinity sites, suggesting that they bound at different sites on the cellulose. A polypeptide, BspCBM17/ CBM28, comprising the tandem CBMs from Ce15A, bound to amorphous cellulose with a significantly higher affinity than the sum of the affinities of CcCBM17 and BspCBM28, indicating cooperativity between the linked CBMs. Ce15A mutants were constructed that were defective in one or both of the CBMs. The mutants differed from the wild-type enzyme in the amounts and sizes of the soluble products produced from amorphous cellulose. This suggests that either the CBMs can modify the action of the catalytic module of Ce15A or that they target the enzyme to areas of the cellulose that differ in susceptibility to hydrolysis.
引用
收藏
页码:6120 / 6127
页数:8
相关论文
共 40 条
[31]   Recognition of cello-oligosaccharides by a family 17 carbohydrate-binding module: An X-ray crystallographic, thermodynamic and mutagenic study [J].
Notenboom, V ;
Boraston, AB ;
Chiu, P ;
Freelove, ACJ ;
Kilburn, DG ;
Rose, DR .
JOURNAL OF MOLECULAR BIOLOGY, 2001, 314 (04) :797-806
[32]  
Sambrook J., 1989, MOL CLONING
[33]   The structural basis for the ligand specificity of family 2 carbohydrate-binding modules [J].
Simpson, PJ ;
Xie, HF ;
Bolam, DN ;
Gilbert, HJ ;
Williamson, MP .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2000, 275 (52) :41137-41142
[34]   A study of the conformation of β-1,4-linked glucose oligomers, cellobiose to cellohexaose, in solution [J].
Sugiyama, H ;
Hisamichi, K ;
Usui, T ;
Sakai, K ;
Ishiyama, J .
JOURNAL OF MOLECULAR STRUCTURE, 2000, 556 (1-3) :173-177
[35]   Hydrophobic surfaces in saccharide chains [J].
Sundari, CS ;
Balasubramanian, D .
PROGRESS IN BIOPHYSICS & MOLECULAR BIOLOGY, 1997, 67 (2-3) :183-216
[36]   CLUSTAL-W - IMPROVING THE SENSITIVITY OF PROGRESSIVE MULTIPLE SEQUENCE ALIGNMENT THROUGH SEQUENCE WEIGHTING, POSITION-SPECIFIC GAP PENALTIES AND WEIGHT MATRIX CHOICE [J].
THOMPSON, JD ;
HIGGINS, DG ;
GIBSON, TJ .
NUCLEIC ACIDS RESEARCH, 1994, 22 (22) :4673-4680
[37]   STUDIES OF THE CELLULOLYTIC SYSTEM OF TRICHODERMA-REESEI QM-9414 - ANALYSIS OF DOMAIN FUNCTION IN 2 CELLOBIOHYDROLASES BY LIMITED PROTEOLYSIS [J].
TOMME, P ;
VANTILBEURGH, H ;
PETTERSSON, G ;
VANDAMME, J ;
VANDEKERCKHOVE, J ;
KNOWLES, J ;
TEERI, T ;
CLAEYSSENS, M .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1988, 170 (03) :575-581
[38]   Interaction of polysaccharides with the N-terminal cellulose-binding domain of Cellulomonas fimi CenC .1. Binding specificity and calorimetric analysis [J].
Tomme, P ;
Creagh, AL ;
Kilburn, DG ;
Haynes, CA .
BIOCHEMISTRY, 1996, 35 (44) :13885-13894
[39]  
Tomme P, 1995, ACS SYM SER, V618, P142
[40]   The binding pattern of two carbohydrate-binding modules of laminarinase Lam16A from Thermotoga neapolitana:: differences in β-glucan binding within family CBM4 [J].
Zverlov, VV ;
Volkov, IY ;
Velikodvorskaya, GA ;
Schwarz, WH .
MICROBIOLOGY-SGM, 2001, 147 :621-629