EzCatDB: D00827
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DB codeD00827
RLCP classification9.1050.192630.8500 : Hydride transfer
8.131.704320.8015 : Isomerization
5.1020.2074300.8301 : Elimination
4.1010.695600.8301 : Addition
8.113.904850.8015 : Isomerization
9.5010.194230.8500 : Hydride transfer
CATH domainDomain 13.40.50.720 : Rossmann fold
Domain 23.90.110.10 : L-2-Hydroxyisocaproate Dehydrogenase; Chain A, domain 2Catalytic domain
E.C.3.2.1.122

CATH domainRelated DB codes (homologues)
3.40.50.720 : Rossmann foldS00543,S00551,S00552,S00553,S00602,S00604,S00605,S00608,S00610,S00625,S00319,S00328,S00329,S00330,S00331,S00332,D00456,D00457,D00458,S00324,S00320,S00325,S00326,S00327,D00459,S00335,S00336,S00334,T00219,S00339,D00513,D00001,D00002,D00003,D00005,D00007,D00008,D00010,D00012,D00017,D00018,D00023,D00027,D00028,D00031,D00032,D00033,D00034,D00035,D00037,D00048,D00071,D00476,D00481,D00482,D00490,D00492,D00494,D00545,D00601,D00603,D00604,D00605,D00615,D00845,D00857,D00858,M00161,M00171,M00210,T00002,T00010,T00011,T00015,T00227,T00247,T00408,T00414,D00262,D00274,D00275,M00035,T00109
3.90.110.10 : L-2-Hydroxyisocaproate Dehydrogenase; Chain A, domain 2D00005,D00008,M00171

Enzyme Name
UniProtKBKEGG

P54716
Protein nameMaltose-6''-phosphate glucosidaseMaltose-6'-phosphate glucosidase
Phospho-alpha-glucosidase
SynonymsEC 3.2.1.122
6-phospho-alpha-D-glucosidase
6-phosphoryl-O-alpha-D-glucopyranosyl:phosphoglucohydrolase
RefSeqNP_388699.1 (Protein)
NC_000964.3 (DNA/RNA sequence)
PfamPF02056 (Glyco_hydro_4)
PF11975 (Glyco_hydro_4C)
[Graphical view]
CAZyGH4 (Glycoside Hydrolase Family)

KEGG pathways
MAP codePathways
MAP00500Starch and sucrose metabolism

UniProtKB:Accession NumberP54716
Entry nameGLVA_BACSU
ActivityMaltose 6'-phosphate + H(2)O = D-glucose + D-glucose 6-phosphate.
SubunitHomotetramer.
Subcellular location
CofactorBinds 1 divalent metal ion such as manganese, iron, cobalt or nickel per subunit.,Binds 1 NAD per subunit.

Compound table: links to PDB-related databases & PoSSuM

CofactorsSubstratesProductsintermediates
KEGG-idC00034C00003C02995C00001C00031C00668I00113I00114I00115I00116I00117
CompoundManganeseNAD+maltose 6'-phosphateH2OD-glucosealpha-D-Glucose 6-phosphateMaltose 3'-dehydro-6'-phosphate2,3-ene-6-phosphate-alpha-D-allopyranosyl-(1->4)-D-glucoseD-glucose-1-deoxy-3-dehydro-6-phosphate-1,2-enealpha-D-allose-2,3-ene-6-phosphatealpha-D-glucose-3-dehydro-6-phosphate
Typeheavy metalamide group,amine group,nucleotidephosphate group/phosphate ion,polysaccharideH2Ocarbohydratecarbohydrate,phosphate group/phosphate ion




ChEBI18291
35154
15846
15703
15377
4167
17665





PubChem23930
5893
439874
962
22247451
5793
439284





                   
1nrhX01UnboundBound:NADUnbound UnboundUnboundUnboundUnboundUnboundUnboundUnbound
1u8xX01UnboundBound:NADUnbound UnboundUnboundUnboundUnboundUnboundUnboundUnbound
1nrhX02Bound:_MNUnboundUnbound UnboundAnalogue:G6PUnboundUnboundUnboundUnboundUnbound
1u8xX02Bound:_MNUnboundUnbound UnboundAnalogue:G6PUnboundUnboundUnboundUnboundUnbound

Active-site residues
resource
Literature [3], [9] & Swiss-prot;P54716
pdbCatalytic residuesCofactor-binding residues
          
1nrhX01GLU 111
 
1u8xX01GLU 111
 
1nrhX02ASP 172;TYR 265
CYS 171;HIS 202 (Manganese binding)
1u8xX02ASP 172;TYR 265
CYS 171;HIS 202 (Manganese binding)

References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[3]Figure 4, Figure 5, p.1626-1627
[8]Figure 1, p.147-149
[9]Figure 1, p.9851
[10]Figure 7, p.535-537

references
[1]
PubMed ID9765262
JournalJ Biol Chem
Year1998
Volume273
Pages27347-56
AuthorsThompson J, Pikis A, Ruvinov SB, Henrissat B, Yamamoto H, Sekiguchi J
TitleThe gene glvA of Bacillus subtilis 168 encodes a metal-requiring, NAD(H)-dependent 6-phospho-alpha-glucosidase. Assignment to family 4 of the glycosylhydrolase superfamily.
[2]
PubMed ID15237973
JournalJ Am Chem Soc
Year2004
Volume126
Pages8354-5
AuthorsYip VL, Varrot A, Davies GJ, Rajan SS, Yang X, Thompson J, Anderson WF, Withers SG
TitleAn unusual mechanism of glycoside hydrolysis involving redox and elimination steps by a family 4 beta-glycosidase from Thermotoga maritima.
[3]
CommentsX-RAY CRYSTALLOGRAPHY (2.05 ANGSTROMS) OF COMPLEX WITH NAD(H) AND ALPHA-D-GLUCOSE-6-PHOSPHATE, REACTION MECHANISM.
PubMed ID15341727
JournalStructure
Year2004
Volume12
Pages1619-29
AuthorsRajan SS, Yang X, Collart F, Yip VL, Withers SG, Varrot A, Thompson J, Davies GJ, Anderson WF
TitleNovel catalytic mechanism of glycoside hydrolysis based on the structure of an NAD+/Mn2+ -dependent phospho-alpha-glucosidase from Bacillus subtilis.
Related PDB1u8x
Related UniProtKBP54716
[4]
PubMed ID16263268
JournalCurr Opin Struct Biol
Year2005
Volume15
Pages637-45
AuthorsDavies GJ, Gloster TM, Henrissat B
TitleRecent structural insights into the expanding world of carbohydrate-active enzymes.
[5]
PubMed ID15670594
JournalJ Mol Biol
Year2005
Volume346
Pages423-35
AuthorsVarrot A, Yip VL, Li Y, Rajan SS, Yang X, Anderson WF, Thompson J, Withers SG, Davies GJ
TitleNAD+ and metal-ion dependent hydrolysis by family 4 glycosidases: structural insight into specificity for phospho-beta-D-glucosides.
[6]
JournalBiocatalysis and Biotransformation
Year2006
Volume24i1-2)
Pages167-176
AuthorsVivian L. Y. Yip; Stephen G. Withers
TitleFamily 4 glycoside hydrolases are special: The first ƒÀ-elimination mechanism amongst glycoside hydrolases
[7]
PubMed ID16401086
JournalBiochemistry
Year2006
Volume45
Pages571-80
AuthorsYip VL, Withers SG
TitleMechanistic analysis of the unusual redox-elimination sequence employed by Thermotoga maritima BglT: a 6-phospho-beta-glucosidase from glycoside hydrolase family 4.
[8]
PubMed ID16495121
JournalCurr Opin Chem Biol
Year2006
Volume10
Pages147-55
AuthorsYip VL, Withers SG
TitleBreakdown of oligosaccharides by the process of elimination.
[9]
CommentsREACTION MECHANISM, CHARACTERIZATION, COFACTORS, SUBSTRATE SPECIFICITY, ENZYME REGULATION, BIOPHYSICOCHEMICAL PROPERTIES.
PubMed ID17676871
JournalBiochemistry
Year2007
Volume46
Pages9840-52
AuthorsYip VL, Thompson J, Withers SG
TitleMechanism of GlvA from Bacillus subtilis: a detailed kinetic analysis of a 6-phospho-alpha-glucosidase from glycoside hydrolase family 4.
[10]
PubMed ID18625333
JournalCurr Opin Chem Biol
Year2008
Volume12
Pages532-8
AuthorsTanner ME
TitleTransient oxidation as a mechanistic strategy in enzymatic catalysis.

comments
This enzyme belongs to glycosidase family-4, with a retaining mechanism.
This glycosidase family-4 is distinct from other glycosidase families in that it requires NAD cofactor and a divalent metal ion for the reaction, and includes both alpha-glycosidase and beta-glycosidase (see [3], [8], [9] and [10]).
According to the literature [3], [9] and [10], this enzyme catalyzes the following reactions:
(A) Hydride transfer from C3 of substrate to nicotinamide of NAD to form 3-keto intermediate (I00113):
(B) Isomerization from 3-keto intermediate to enediolate intermediate (I00114):
(C) Elimination of D-glucose from C1 atom of the enediolate intermediate, forming enone intermediate (I00115):
(D) Addition of water to C1 atom of the enone intermediate, forming enediolate intermediate (I00116):
(E) Isomerization from enediolate intermediate to 3-keto intermediate (I00117):
(F) Hydride transfer from nicotinamide of NADH to C3 atom of the 3-keto intermediate:
The detailed mechanism seems to be as follows:
(A) Hydride transfer from C3 of substrate to nicotinamide of NAD to form 3-keto intermediate (I00113):
(A0) Manganese ion, which is bound to Cys171, His202 and O7 atom of nicotinamide of NAD, binds O3 and O2 hydroxyl groups, lowering the pKa of O3 hydroxyl group. This facilitates the deprotonation of O3 hydroxyl group.
(A1) Hydride transfer from C3 atom of substrate to nicotinamide of NAD occurs, at the same time as the deprotonation of O3 hydroxyl group, forming 3-keto intermediate (I00113).
(B) Isomerization from 3-keto intermediate to enediolate intermediate (I00114):
(B0) Oxidation of C3 atom acidifies C2 atom, which facilitates the deprotonation of C2 atom. Meanwhile, the manganese ion also facilitate the deprotonation of C2 atom by polarizing carbonyl group at C3. On the other hand, Glu111 modulates the activity of Tyr265 through a water molecule.
(B1) Tyr265 acts as a general base to deprotonate C2 atom, forming an edediolate intermediate (I00114).
(C) Elimination of D-glucose from C1 atom of the enediolate intermediate, forming enone intermediate (I00115):
(C0) The enediolate intermediate, bound to the manganese ion, is reactive.
(C1) Asp172 acts as a general acid to protonate the oxygen atom of the eliminated D-glucose group. This elimination forms an enone intermediate (I00115).
(D) Addition of water to C1 atom of the enone intermediate, forming enediolate intermediate (I00116):
(D0) The enone intermediate (or alpha,beta-unsaturated intermediate), bound to the manganese ion, is reactive for addition at C1.
(D1) Asp172 acts as a general base to deprotonate a water molecule.
(D2) The activated water attacks on the C1 atom of the enone intermediate, forming an enediolate intermediate (I00116).
(E) Isomerization from enediolate intermediate to 3-keto intermediate (I00117):
(E0) O2 and O3 atoms of the enediolate are bound to the manganese ion. On the other hand, Glu111 modulates the activity of Tyr265 through a water molecule.
(E1) Tyr265 acts as a general acid to protonate C2 atom, forming 3-keto intermediate (I00117).
(F) Hydride transfer from nicotinamide of NADH to C3 atom of the 3-keto intermediate:
(F0) Manganese ion, which is bound to Cys171, His202 and O7 atom of nicotinamide of NAD, binds O3 and O2 atoms of the enediolate intermediate.
(F1) Hydride transfer from nicotinamide of NADH to C3 atom of the intermediate occurs, at the same time as the protonation to O3 atom, forming final product.

createdupdated
2009-12-082011-12-27


Copyright: Nozomi Nagano, JST & CBRC-AIST
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Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2005 - March 2006)
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Funded by BIRD/Japan Science and Technology Corporation (JST) (September 2005 - September 2008)
Funded by BIRD/Japan Science and Technology Corporation (JST) (October 2007 - September 2010)
Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2011 - March 2012)
Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2012 - March 2013)
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