DB code: S00912

RLCP classification 1.30.35880.983 : Hydrolysis
CATH domain 3.20.20.80 : TIM Barrel Catalytic domain
E.C. 3.2.1.89
CSA
M-CSA
MACiE

CATH domain Related DB codes (homologues)
3.20.20.80 : TIM Barrel S00202 S00210 S00748 S00906 S00907 S00911 S00915 M00134 M00160 D00479 S00204 S00205 S00206 S00207 S00203 S00208 S00209 S00211 S00213 S00214 M00113 T00307 D00165 D00166 D00169 D00176 D00501 D00502 D00503 D00844 D00861 D00864 M00026 M00112 M00193 M00346 T00057 T00062 T00063 T00066 T00067

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq CAZy Pfam
Q65CX5 Arabinogalactan endo-1,4-beta-galactosidase
EC 3.2.1.89
Endo-1,4-beta-galactanase
Galactanase
YP_006715543.1 (Protein)
NC_006322.1 (DNA/RNA sequence)
YP_081349.1 (Protein)
NC_006270.3 (DNA/RNA sequence)
GH53 (Glycoside Hydrolase Family 53)
PF07745 (Glyco_hydro_53)
[Graphical View]

KEGG enzyme name
Arabinogalactan endo-1,4-beta-galactosidase
Endo-1,4-beta-galactanase
Galactanase
Arabinogalactanase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
Q65CX5 GANA_BACLD Endohydrolysis of 1,4-beta-D-galactosidic linkages in arabinogalactans. Binds 1 calcium ion per subunit.

KEGG Pathways
Map code Pathways E.C.

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00076 L00048 L00049 C05796 C00001 L00048 L00049 C05796 C03611 I00121
E.C.
Compound Calcium Rhamnogalacturonan I Arabinogalactan type I Galactan H2O Rhamnogalacturonan I Arabinogalactan Galactan Galactose oligosaccharide Peptidyl-Glu-galactan
Type divalent metal (Ca2+, Mg2+) carboxyl group,polysaccharide polysaccharide polysaccharide H2O carboxyl group,polysaccharide polysaccharide polysaccharide polysaccharide
ChEBI 29108
15377
PubChem 271
22247451
962
1r8lA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1r8lB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Unbound Unbound Unbound
1ur0A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Unbound Unbound
1ur0B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Unbound Unbound
1ur4A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B2G Unbound Unbound
1ur4B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B2G Unbound Unbound
2ccrA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Bound:B2G Unbound
2ccrB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Bound:B2G Unbound
2gftA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:GAL-GAL-GAL Unbound Unbound
2gftB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:GAL-GAL-GAL Unbound Unbound
2j74A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Bound:B2G Unbound
2j74B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_CA Unbound Unbound Unbound Unbound Unbound Bound:B4G Bound:B2G Unbound

Reference for Active-site residues
resource references E.C.
Literature [6], [8], [9], [11], Swiss-prot;Q65CX5, P48842, P83691 & P83692

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1r8lA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
1r8lB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
1ur0A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
1ur0B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
1ur4A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
1ur4B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
2ccrA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
2ccrB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
2gftA00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234; ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding) mutant E236A
2gftB00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 166;TYR 234; ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding) mutant E236A
2j74A00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)
2j74B00 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ARG 66;ASN 164;GLU 165;TYR 234;GLU 263 ASP 272;ASP 274;HIS 276;ASN 278;SER 367;ASP 370 (Calcium binding)

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[6]
FIGURE 3, p.15138-15141
[8]
Figure 5
[9]
p.109
[11]
p.978

References
[1]
Resource
Comments
Medline ID
PubMed ID 823153
Journal J Biol Chem
Year 1976
Volume 251
Pages 5904-10
Authors Labavitch JM, Freeman LE, Albersheim P
Title Structure of plant cell walls. Purification and characterization of a beta-1,4-galactanase which degrades a structural component of the primary cell walls of dicots.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID
Journal Carbohydr Polym
Year 1991
Volume 16
Pages 167-87
Authors van de Visa JW, Searle-van Leeuwena MJF, Silihaa HA, Kormelinka FJM, Voragena AGJ
Title Purification and characterization of Endo-1,4-beta-D-galactanases from Aspergillus niger and Aspergillus aculeatus: Use in combination with arabinanases from Aspergillus niger in enzymic conversion of potato arabinogalactan
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID 7712292
Journal Curr Opin Struct Biol
Year 1994
Volume 4
Pages 885-92
Authors McCarter JD, Withers SG
Title Mechanisms of enzymatic glycoside hydrolysis.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 8535779
Journal Structure
Year 1995
Volume 3
Pages 853-9
Authors Davies G, Henrissat B
Title Structures and mechanisms of glycosyl hydrolases.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 9398278
Journal Biochemistry
Year 1997
Volume 36
Pages 15489-500
Authors Braithwaite KL, Barna T, Spurway TD, Charnock SJ, Black GW, Hughes N, Lakey JH, Virden R, Hazlewood GP, Henrissat B, Gilbert HJ
Title Evidence that galactanase A from Pseudomonas fluorescens subspecies cellulosa is a retaining family 53 glycosyl hydrolase in which E161 and E270 are the catalytic residues.
Related PDB
Related UniProtKB
[6]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.8 ANGSTROMS).
Medline ID
PubMed ID 12484750
Journal Biochemistry
Year 2002
Volume 41
Pages 15135-43
Authors Ryttersgaard C, Lo Leggio L, Coutinho PM, Henrissat B, Larsen S
Title Aspergillus aculeatus beta-1,4-galactanase: substrate recognition and relations to other glycoside hydrolases in clan GH-A.
Related PDB 1fhl 1fob
Related UniProtKB P48842
[7]
Resource
Comments
Medline ID
PubMed ID
Journal Carbohydr Polym
Year 2003
Volume 53
Pages 155-68
Authors Luonteria E, Laineb C, Uusitaloa S, Telemanc A, Siika-ahoa M, Tenkanen M
Title Purification and characterization of Aspergillus beta-D-galactanases acting on beta-1,4- and beta-1,3/6-linked arabinogalactans
Related PDB
Related UniProtKB
[8]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.6 ANGSTROMS), CATALYTIC ACTIVITY, GLYCOSYLATION AT ASN-111.
Medline ID
PubMed ID 12761390
Journal Protein Sci
Year 2003
Volume 12
Pages 1195-204
Authors Le Nours J, Ryttersgaard C, Lo Leggio L, Ostergaard PR, Borchert TV, Christensen LL, Larsen S
Title Structure of two fungal beta-1,4-galactanases: searching for the basis for temperature and pH optimum.
Related PDB 1hjq 1hjs 1hju
Related UniProtKB P83691 P83692
[9]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.2 ANGSTROMS) OF 28-424 IN COMPLEX WITH CALCIUM IONS AND SUBSTRATE ANALOGS, COFACTOR.
Medline ID
PubMed ID 15312766
Journal J Mol Biol
Year 2004
Volume 341
Pages 107-17
Authors Ryttersgaard C, Le Nours J, Lo Leggio L, Jorgensen CT, Christensen LL, Bjornvad M, Larsen S
Title The structure of endo-beta-1,4-galactanase from Bacillus licheniformis in complex with two oligosaccharide products.
Related PDB 1r8l 1ur0 1ur4
Related UniProtKB Q65CX5
[10]
Resource
Comments
Medline ID
PubMed ID 16151143
Journal Appl Environ Microbiol
Year 2005
Volume 71
Pages 5501-10
Authors Hinz SW, Pastink MI, van den Broek LA, Vincken JP, Voragen AG
Title Bifidobacterium longum endogalactanase liberates galactotriose from type I galactans.
Related PDB
Related UniProtKB
[11]
Resource
Comments
Medline ID
PubMed ID 19089956
Journal Proteins
Year 2009
Volume 75
Pages 977-89
Authors Le Nours J, De Maria L, Welner D, Jorgensen CT, Christensen LL, Borchert TV, Larsen S, Lo Leggio L
Title Investigating the binding of beta-1,4-galactan to Bacillus licheniformis beta-1,4-galactanase by crystallography and computational modeling.
Related PDB 2ccr 2j74
Related UniProtKB Q65CX5

Comments
This enzyme belongs to glycosidase family-58, with a retaining mechanism.
This enzyme is homologous to a counterpart enzyme from fungi (S00748 in EzCatDB).
Although this enzyme binds calcium ion, it is not involved in catalysis. The reactiom mechanism must be similar to that of the homologous enzyme.
According to the literature [6], the reaction mechanism proceeds as follows:
(0) Arg66 and Tyr234 modulate the pKa of catalytic nucleophile, Glu263.
(1) Glu165 acts as a general acid to protonate the leaving oxygen, O4 atom at subsite +1, forming an oxocarbenium-ion-like transition-state. The transition-state is stabilized by Asn164 and Tyr234 through hydrogen bonding to the oxygen atoms at subsite -1. (SN1-like reaction)
(2) Glu263 makes a nucleophilic attack on C1 atom of substrate, galactan, to form a covalent intermediate.
(3) Glu165 acts as a general base to deprotonate a water molecule, to activate it.
(4) The activated water makes a nucleophilic attack on the C1 atom of the covalent intermediate. Thus, the substitution reaction completes.

Created Updated
2010-04-08 2018-04-23