4QLL: Crystal Structure Of Rice Bglu1 E176q/y341a/q187a Mutant Complexed With Cellotetraose

Citation:
Abstract
Rice BGlu1 (Os3BGlu7) is a glycoside hydrolase family 1 beta-glucosidase that hydrolyzes cellooligosaccharides with increasing efficiency as the degree of polymerization (DP) increases from 2 to 6, indicating six subsites for glucosyl residue binding. Five subsites have been identified in X-ray crystal structures of cellooligosaccharide complexes with its E176Q acid-base and E386G nucleophile mutants. X-ray crystal structures indicate that cellotetraose binds in a similar mode in BGlu1 E176Q and E386G, but in a different mode in the BGlu1 E386G/Y341A variant, in which glucosyl residue 4 (Glc4) interacts with Q187 instead of the eliminated phenolic group of Y341. Here, we found that the Q187A mutation has little effect on BGlu1 cellooligosaccharide hydrolysis activity or oligosaccharide binding in BGlu1 E176Q, and only slight effects on BGlu1 E386G glycosynthase activity. X-ray crystal structures showed that cellotetraose binds in a different position in BGlu1 E176Q/Y341A, in which it interacts directly with R178 and W337, and the Q187A mutation had little effect on cellotetraose binding. Mutations of R178 and W337 to A had significant and nonadditive effects on oligosaccharide hydrolysis by BGlu1, pNPGlc cleavage and cellooligosaccharide inhibition of BGlu1 E176Q and BGlu1 E386G glycosynthase activity. Hydrolysis activity was partially rescued by Y341 for longer substrates, suggesting stacking of Glc4 on Y341 stabilizes binding of cellooligosaccharides in the optimal position for hydrolysis. This analysis indicates that complex interactions between active site cleft residues modulate substrate binding and hydrolysis.
PDB ID: 4QLLDownload
MMDB ID: 130053
PDB Deposition Date: 2014/6/12
Updated in MMDB: 2015/06
Experimental Method:
x-ray diffraction
Resolution: 1.85  Å
Source Organism:
Similar Structures:
Biological Unit for 4QLL: monomeric; determined by author and by software (PISA)
Molecular Components in 4QLL
Label Count Molecule
Protein (1 molecule)
1
Beta-glucosidase 7(Gene symbol: LOC4333841)
Molecule annotation
Chemicals (5 molecules)
1
2
2
1
3
1
4
1
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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