Crystal structure of mannanase 26A from Pseudomonas cellulosa and analysis of residues involved in substrate binding

被引:81
作者
Hogg, D
Woo, EJ
Bolam, DN
McKie, VA
Gilbert, HJ
Pickersgill, RW
机构
[1] Queen Mary Univ London, Sch Biol Sci, London E1 4NS, England
[2] Newcastle Univ, Dept Biol & Nutr Sci, Newcastle Upon Tyne NE1 7RU, Tyne & Wear, England
关键词
D O I
10.1074/jbc.M010290200
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
The crystal structure of Pseudomonas cellulosa mannanase 26A has been solved by multiple isomorphous replacement and refined at 1.85 Angstrom resolution to an R-factor of 0.182 (R-free = 0.211). The enzyme comprises (beta/alpha)(8)-barrel architecture with two catalytic glutamates at the ends of beta -strands 4 and 7 in precisely the same location as the corresponding glutamates in other 4/7-superfamily glycoside hydrolase enzymes (clan GH-A glycoside hydrolases). The family 26 glycoside hydrolases are therefore members of clan GH-A. Functional analyses of mannanase 26A, informed by the crystal structure of the enzyme, provided important insights into the role of residues close to the catalytic glutamates. These data showed that Trp-360 played a critical role in binding substrate at the -1 subsite, whereas Tyr-285 was important to the function of the nucleophile catalyst. His-211 in mannanase 26A does not have the same function as the equivalent asparagine in the other GH-A enzymes. The data also suggest that Trp-217 and Trp-162 are important for the activity of mannanase 26RA against mannooligosaccharides but are less important for activity against polysaccharides.
引用
收藏
页码:31186 / 31192
页数:7
相关论文
共 34 条
[1]   Influence of the aglycone region of the substrate binding cleft of Pseudomonas xylanase 10A on catalysis [J].
Armand, S ;
Andrews, SR ;
Charnock, SJ ;
Gilbert, HJ .
BIOCHEMISTRY, 2001, 40 (25) :7404-7409
[2]   THE CCP4 SUITE - PROGRAMS FOR PROTEIN CRYSTALLOGRAPHY [J].
BAILEY, S .
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY, 1994, 50 :760-763
[3]   STRUCTURE OF CHICKEN MUSCLE TRIOSE PHOSPHATE ISOMERASE DETERMINED CRYSTALLOGRAPHICALLY AT 2.5A RESOLUTION USING AMINO-ACID SEQUENCE DATA [J].
BANNER, DW ;
BLOOMER, AC ;
PETSKO, GA ;
PHILLIPS, DC ;
POGSON, CI ;
WILSON, IA ;
CORRAN, PH ;
FURTH, AJ ;
MILMAN, JD ;
OFFORD, RE ;
PRIDDLE, JD ;
WALEY, SG .
NATURE, 1975, 255 (5510) :609-614
[4]   Mannanase A from Pseudomonas fluorescens ssp cellulosa is a retaining glycosyl hydrolase in which E212 and E320 are the putative catalytic residues [J].
Bolam, DN ;
Hughes, N ;
Virden, R ;
Lakey, JH ;
Hazlewood, GP ;
Henrissat, B ;
Braithwaite, KL ;
Gilbert, HJ .
BIOCHEMISTRY, 1996, 35 (50) :16195-16204
[5]   A NON-MODULAR ENDO-BETA-1,4-MANNANASE FROM PSEUDOMONAS-FLUORESCENS SUBSPECIES CELLULOSA [J].
BRAITHWAITE, KL ;
BLACK, GW ;
HAZLEWOOD, GP ;
ALI, BRS ;
GILBERT, HJ .
BIOCHEMICAL JOURNAL, 1995, 305 :1005-1010
[6]   The topology of the substrate binding clefts of glycosyl hydrolase family 10 xylanases are not conserved [J].
Charnock, SJ ;
Spurway, TD ;
Xie, HF ;
Beylot, MH ;
Virden, R ;
Warren, RAJ ;
Hazlewood, GP ;
Gilbert, HJ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1998, 273 (48) :32187-32199
[7]   Key residues in subsite F play a critical role in the activity of Pseudomonas fluorescens subspecies cellulosa xylanase A against xylooligosaccharides but not against highly polymeric substrates such as xylan [J].
Charnock, SJ ;
Lakey, JH ;
Virden, R ;
Hughes, N ;
Sinnott, ML ;
Hazlewood, GP ;
Pickersgill, R ;
Gilbert, HJ .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1997, 272 (05) :2942-2951
[8]   Miscellaneous algorithms for density modification [J].
Cowtan, K ;
Main, P .
ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY, 1998, 54 :487-493
[9]   Structure of the Bacillus agaradherans family 5 endoglucanase at 1.6 Å and its cellobiose complex at 2.0 Å resolution [J].
Davies, GJ ;
Dauter, M ;
Brzozowski, AM ;
Bjornvad, ME ;
Andersen, KV ;
Schülein, M .
BIOCHEMISTRY, 1998, 37 (07) :1926-1932
[10]   Leaving no element of doubt: analysis of proteins using microPIXE [J].
Garman, E .
STRUCTURE WITH FOLDING & DESIGN, 1999, 7 (12) :R291-R299