DB code: D00175

RLCP classification	1.30.36040.982 : Hydrolysis
CATH domain	3.20.20.300 : TIM Barrel	Catalytic domain
CATH domain	3.40.50.1700 : Rossmann fold	Catalytic domain
E.C.	3.2.1.-
CSA
M-CSA
MACiE

CATH domain	Related DB codes (homologues)

Uniprot Enzyme Name
UniprotKB	Protein name	Synonyms	CAZy	Pfam
Q9XEI3		Beta-D-glucan exohydrolase isoenzyme ExoI	GH3 (Glycoside Hydrolase Family 3)	PF00933 (Glyco_hydro_3) PF01915 (Glyco_hydro_3_C) [Graphical View]

KEGG enzyme name

UniprotKB: Accession Number	Entry name	Activity	Subunit	Subcellular location	Cofactor
Q9XEI3	Q9XEI3_HORVD

KEGG Pathways
Map code	Pathways	E.C.

Compound table
	Substrates		Products		Intermediates
KEGG-id	C00965	C00001	C00221	C00965
E.C.
Compound	1,3-beta-D-Glucan	H2O	beta-D-Glucose	1,3-beta-D-Glucan	Transition-state in glycosylation	Glycosyl-enzyme intermediate	Transition-state in deglycosylation
Type	polysaccharide	H2O	carbohydrate	polysaccharide
ChEBI		15377 15377	15903 15903
PubChem		22247451 962 22247451 962	64689 64689

1ex1A01	Unbound		Analogue:GLC	Unbound	Unbound	Unbound	Unbound
1ieqA01	Unbound		Bound:GLC	Unbound	Unbound	Unbound	Unbound
1ievA01	Unbound		Unbound	Unbound	Unbound	Intermediate-analogue:INS	Unbound
1iewA01	Unbound		Unbound	Unbound	Unbound	Intermediate-analogue:G2F	Unbound
1iexA01	Analogue:TCB		Unbound	Unbound	Unbound	Unbound	Unbound
1j8vA01	Analogue:LAM		Unbound	Unbound	Unbound	Unbound	Unbound
1lq2A01	Unbound		Unbound	Unbound	Transition-state-analogue:IDD	Unbound	Unbound
1x38A01	Unbound		Unbound	Unbound	Transition-state-analogue:IDD	Unbound	Unbound
1x39A01	Unbound		Unbound	Unbound	Transition-state-analogue:IDE	Unbound	Unbound
1ex1A02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1ieqA02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1ievA02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1iewA02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1iexA02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1j8vA02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1lq2A02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1x38A02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound
1x39A02	Unbound		Unbound	Unbound	Unbound	Unbound	Unbound

Reference for Active-site residues
resource	references	E.C.
PDB;1ex1 & literature [3], [6]

Active-site residues
PDB	Catalytic residues	Cofactor-binding residues	Modified residues	Main-chain involved in catalysis	Comment

1ex1A01	ARG 158;ASP 285
1ieqA01	ARG 158;ASP 285
1ievA01	ARG 158;ASP 285
1iewA01	ARG 158;ASP 285
1iexA01	ARG 158;ASP 285
1j8vA01	ARG 158;ASP 285
1lq2A01	ARG 158;ASP 285
1x38A01	ARG 158;ASP 285
1x39A01	ARG 158;ASP 285
1ex1A02	TRP 430;TRP 434;GLU 491
1ieqA02	TRP 430;TRP 434;GLU 491
1ievA02	TRP 430;TRP 434;GLU 491
1iewA02	TRP 430;TRP 434;GLU 491
1iexA02	TRP 430;TRP 434;GLU 491
1j8vA02	TRP 430;TRP 434;GLU 491
1lq2A02	TRP 430;TRP 434;GLU 491
1x38A02	TRP 430;TRP 434;GLU 491
1x39A02	TRP 430;TRP 434;GLU 491

References for Catalytic Mechanism
References	Sections	No. of steps in catalysis
[3]	p.186-187
[4]	p.263-266
[5]	Fig.3, p.85-86
[6]	Fig.8, p.1013-1014
[9]	p.4976-4979
[10]	p.16536-16538

References
[1]
Resource
Comments
Medline ID
PubMed ID	8617814
Journal	J Biol Chem
Year	1996
Volume	271
Pages	5277-86
Authors	Hrmova M, Harvey AJ, Wang J, Shirley NJ, Jones GP, Stone BA, Hoj PB, Fincher GB
Title	Barley beta-D-glucan exohydrolases with beta-D-glucosidase activity. Purification, characterization, and determination of primary structure from a cDNA clone.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID
Journal	Carbohydr Res
Year	1998
Volume	305
Pages	209-21
Authors	Hrmova M, Fincher GB
Title	Barley beta-D-glucan exohydrolases. Substrate specificity and kinetic properties.
Related PDB
Related UniProtKB
[3]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF 26-630.
Medline ID
PubMed ID	10368285
Journal	Structure Fold Des
Year	1999
Volume	7
Pages	179-90
Authors	Varghese JN, Hrmova M, Fincher GB
Title	Three-dimensional structure of a barley beta-D-glucan exohydrolase, a family 3 glycosyl hydrolase.
Related PDB	1ex1
Related UniProtKB	Q9XEI3
[4]
Resource
Comments
Medline ID
PubMed ID	10966578
Journal	Proteins
Year	2000
Volume	41
Pages	257-69
Authors	Harvey AJ, Hrmova M, De Gori R, Varghese JN, Fincher GB
Title	Comparative modeling of the three-dimensional structures of family 3 glycoside hydrolases.
Related PDB
Related UniProtKB
[5]
Resource
Comments	structure-function relationships (review)
Medline ID
PubMed ID	11554481
Journal	Plant Mol Biol
Year	2001
Volume	47
Pages	73-91
Authors	Hrmova M, Fincher GB
Title	Structure-function relationships of beta-D-glucan endo- and exohydrolases from higher plants.
Related PDB
Related UniProtKB
[6]
Resource
Comments	X-ray crystallography
Medline ID
PubMed ID	11709165
Journal	Structure
Year	2001
Volume	9
Pages	1005-16
Authors	Hrmova M, Varghese JN, De Gori R, Smith BJ, Driguez H, Fincher GB
Title	Catalytic mechanisms and reaction intermediates along the hydrolytic pathway of a plant beta-D-glucan glucohydrolase.
Related PDB	1ieq 1iev 1iew 1iex
Related UniProtKB
[7]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (2.4 ANGSTROMS) OF 26-630.
Medline ID
PubMed ID	12034895
Journal	Plant Cell
Year	2002
Volume	14
Pages	1033-52
Authors	Hrmova M, De Gori R, Smith BJ, Fairweather JK, Driguez H, Varghese JN, Fincher GB
Title	Structural basis for broad substrate specificity in higher plant beta-D-glucan glucohydrolases.
Related PDB	1j8v
Related UniProtKB	Q9XEI3
[8]
Resource
Comments
Medline ID
PubMed ID	12932786
Journal	J Mol Graph Model
Year	2003
Volume	22
Pages	151-9
Authors	Smith BJ
Title	Can thioglycosides imitate the oxonium intermediate in glycosyl hydrolases?
Related PDB
Related UniProtKB
[9]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (2.7 ANGSTROMS) OF 26-627.
Medline ID
PubMed ID	14597633
Journal	J Biol Chem
Year	2004
Volume	279
Pages	4970-80
Authors	Hrmova M, De Gori R, Smith BJ, Vasella A, Varghese JN, Fincher GB
Title	Three-dimensional structure of the barley beta-D-glucan glucohydrolase in complex with a transition state mimic.
Related PDB	1lq2
Related UniProtKB	Q9XEI3
[10]
Resource
Comments	X-ray crystallography
Medline ID
PubMed ID	16342944
Journal	Biochemistry
Year	2005
Volume	44
Pages	16529-39
Authors	Hrmova M, Streltsov VA, Smith BJ, Vasella A, Varghese JN, Fincher GB
Title	Structural rationale for low-nanomolar binding of transition state mimics to a family GH3 beta-D-glucan glucohydrolase from barley.
Related PDB	1x38 1x39
Related UniProtKB

Comments
This enzyme belongs to the glycosidase family-3, with a retaining mechanism. According to the literature [1], although the reactions by this enzyme are similar to those by (1,3)-beta-glucan exohydrolases that have been classified in E.C. 3.2.1.58, this enzyme can hydrolyze substrates with linkages other than those in the (1,3)-position. Furthermore, whilst the glycosidases that are classified in the E.C. 3.2.1.58 must have an inverting mechanism, releaseing alpha-D-Glucose, this enzyme has got a retaining mechanism, releasing beta-D-glucose. Thus, the E.C. number of this enzyme is not completely assigned yet. According to the literature [2] & [6], the reaction of this enzyme proceeds as follows: (1) Trp430 and Trp434 may act as modulator by increasing the pKa of Glu491, whereas Arg158 may decrease the pKa of Glu491 as a modulator. (2) Glu491 acts as a general acid to protonate the leaving oxygen atom, leading to the formation of an oxocarbonium ion-like transition-state. Here, Asp285 may stabilize the positive charge on the C1 carbon of the oxocarbonium ion-like transition-state. (3) Asp285 makes a nucleophilic attack on the C1 carbon, forming a glycosyl-enzyme intermediate. (4) Glu491 acts as a general base to activate a water molecule. (5) The activated water molecule makes a nucleophilic attack on the C1 carbon of the intermediate, completing the hydrolysis reaction through the formation of an oxocarbonium ion-like transition-state.

Comments

This enzyme belongs to the glycosidase family-3, with a retaining mechanism.
According to the literature [1], although the reactions by this enzyme are similar to those by (1,3)-beta-glucan exohydrolases that have been classified in E.C. 3.2.1.58, this enzyme can hydrolyze substrates with linkages other than those in the (1,3)-position.
Furthermore, whilst the glycosidases that are classified in the E.C. 3.2.1.58 must have an inverting mechanism, releaseing alpha-D-Glucose, this enzyme has got a retaining mechanism, releasing beta-D-glucose. Thus, the E.C. number of this enzyme is not completely assigned yet.
According to the literature [2] & [6], the reaction of this enzyme proceeds as follows:
(1) Trp430 and Trp434 may act as modulator by increasing the pKa of Glu491, whereas Arg158 may decrease the pKa of Glu491 as a modulator.
(2) Glu491 acts as a general acid to protonate the leaving oxygen atom, leading to the formation of an oxocarbonium ion-like transition-state. Here, Asp285 may stabilize the positive charge on the C1 carbon of the oxocarbonium ion-like transition-state.
(3) Asp285 makes a nucleophilic attack on the C1 carbon, forming a glycosyl-enzyme intermediate.
(4) Glu491 acts as a general base to activate a water molecule.
(5) The activated water molecule makes a nucleophilic attack on the C1 carbon of the intermediate, completing the hydrolysis reaction through the formation of an oxocarbonium ion-like transition-state.

Created	Updated
2004-05-07	2009-02-26