DB code: D00665

RLCP classification	1.30.35890.994 : Hydrolysis
CATH domain	3.20.20.70 : TIM Barrel	Catalytic domain
CATH domain	2.60.40.1180 : Immunoglobulin-like
E.C.	3.2.1.49
CSA
M-CSA
MACiE

CATH domain	Related DB codes (homologues)
2.60.40.1180 : Immunoglobulin-like	M00113 T00307 D00165 D00176 D00664 D00863 D00864 M00112 M00193 M00314 T00057 T00062 T00067
3.20.20.70 : TIM Barrel	S00215 S00217 S00218 S00219 S00532 S00198 S00220 S00745 S00537 S00538 S00539 S00826 S00841 S00235 S00239 S00240 S00243 S00244 S00199 S00200 S00201 S00221 S00222 S00847 S00224 S00225 S00226 D00014 D00029 M00141 T00015 T00239 D00664 D00804 D00863 T00089

Uniprot Enzyme Name
UniprotKB	Protein name	Synonyms	CAZy	Pfam	RefSeq
Q90744	Alpha-N-acetylgalactosaminidase	EC 3.2.1.49 Alpha-galactosidase B	GH27 (Glycoside Hydrolase Family 27)	PF02065 (Melibiase) [Graphical View]
P17050	Alpha-N-acetylgalactosaminidase	EC 3.2.1.49 Alpha-galactosidase B	GH27 (Glycoside Hydrolase Family 27)	PF02065 (Melibiase) [Graphical View]	NP_000253.1 (Protein) NM_000262.2 (DNA/RNA sequence)

KEGG enzyme name
Alpha-N-acetylgalactosaminidase Alpha-acetylgalactosaminidase N-acetyl-alpha-D-galactosaminidase N-acetyl-alpha-galactosaminidase

UniprotKB: Accession Number	Entry name	Activity	Subunit	Subcellular location	Cofactor
Q90744	NAGAB_CHICK	Hydrolysis of terminal non-reducing N-acetyl-D-galactosamine residues in N-acetyl-alpha-D-galactosaminides.	Homodimer.	Lysosome (By similarity).
P17050	NAGAB_HUMAN	Hydrolysis of terminal non-reducing N-acetyl-D-galactosamine residues in N-acetyl-alpha-D-galactosaminides.	Homodimer.	Lysosome.

KEGG Pathways
Map code	Pathways	E.C.
MAP00603	Glycosphingolipid biosynthesis - globoseries

Compound table
	Substrates		Products		Intermediates
KEGG-id	C04134	C00001	C04134	L00046	I00063
E.C.
Compound	N-Acetyl-alpha-D-galactosaminide	H2O	N-Acetyl-alpha-D-galactosaminide	N-Acetyl-alpha-D-galactosamine	Peptidyl-ASP-beta-N-acetylgalactosaminie
Type	amide group,polysaccharide	H2O	amide group,polysaccharide	amide group,carbohydrate
ChEBI		15377 15377		40356 40356
PubChem		22247451 962 22247451 962		84265 84265

1ktbA01	Unbound		Unbound	Unbound	Unbound
1ktcA01	Unbound		Unbound	Bound:NGA	Unbound
3h53A01	Unbound		Unbound	Unbound	Unbound
3h53B01	Unbound		Unbound	Unbound	Unbound
3h54A01	Unbound		Unbound	Bound:A2G	Unbound
3h54B01	Unbound		Unbound	Bound:A2G	Unbound
3h55A01	Unbound		Analogue:GLA_1500	Analogue:GLA_1000	Unbound
3h55B01	Unbound		Analogue:GLA_2500	Analogue:GLA_2000	Unbound
3iguA01	Unbound		Unbound	Unbound	Intermediate-analogue:7JZ
3iguB01	Unbound		Unbound	Unbound	Intermediate-analogue:7JZ
1ktbA02	Unbound		Unbound	Unbound	Unbound
1ktcA02	Unbound		Unbound	Unbound	Unbound
3h53A02	Unbound		Unbound	Unbound	Unbound
3h53B02	Unbound		Unbound	Unbound	Unbound
3h54A02	Unbound		Unbound	Unbound	Unbound
3h54B02	Unbound		Unbound	Unbound	Unbound
3h55A02	Unbound		Unbound	Unbound	Unbound
3h55B02	Unbound		Unbound	Unbound	Unbound
3iguA02	Unbound		Unbound	Unbound	Unbound
3iguB02	Unbound		Unbound	Unbound	Unbound

Reference for Active-site residues
resource	references	E.C.
literature [4], [5]

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

1ktbA01	ASP 61;TYR 103;ASP 140;ARG 197;ASP 201
1ktcA01	ASP 61;TYR 103;ASP 140;ARG 197;ASP 201
3h53A01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3h53B01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3h54A01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3h54B01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3h55A01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3h55B01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3iguA01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
3iguB01	ASP 78;TYR 119;ASP 156;ARG 213;ASP 217
1ktbA02
1ktcA02
3h53A02
3h53B02
3h54A02
3h54B02
3h55A02
3h55B02
3iguA02
3iguB02

References for Catalytic Mechanism
References	Sections	No. of steps in catalysis
[4]	Figure 4
[5]	Fig.6

References
[1]
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
[2]
Resource
Comments
Medline ID
PubMed ID	10933800
Journal	Biochemistry
Year	2000
Volume	39
Pages	9826-36
Authors	Hart DO, He S, Chany CJ 2nd, Withers SG, Sims PF, Sinnott ML, Brumer H 3rd
Title	Identification of Asp-130 as the catalytic nucleophile in the main alpha-galactosidase from Phanerochaete chrysosporium, a family 27 glycosyl hydrolase.
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID	11128583
Journal	Carbohydr Res
Year	2000
Volume	329
Pages	539-47
Authors	Ly HD, Howard S, Shum K, He S, Zhu A, Withers SG
Title	The synthesis, testing and use of 5-fluoro-alpha-D-galactosyl fluoride to trap an intermediate on green coffee bean alpha-galactosidase and identify the catalytic nucleophile.
Related PDB
Related UniProtKB
[4]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (1.9 ANGSTROMS) IN COMPLEX WITH N-ACETYLGALACTOSAMINE, GLYCOSYLATION AT ASN-161; ASN-185 AND ASN-369, SUBUNIT, DISULFIDE BONDS.
Medline ID
PubMed ID	12005440
Journal	Structure
Year	2002
Volume	10
Pages	425-34
Authors	Garman SC, Hannick L, Zhu A, Garboczi DN
Title	The 1.9 A structure of alpha-N-acetylgalactosaminidase: molecular basis of glycosidase deficiency diseases.
Related PDB	1ktb 1ktc
Related UniProtKB	Q90744
[5]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (1.9 ANGSTROMS) OF 18-411 IN COMPLEXES WITH N-ACETYLGALACTOSAMINE AND GALACTOSE, DISULFIDE BONDS, GLYCOSYLATION AT ASN-124; ASN-177 AND ASN-385, SUBUNIT, CATALYTIC ACTIVITY, MUTAGENESIS OF ASN-201.
Medline ID
PubMed ID	19683538
Journal	J Mol Biol
Year	2009
Volume	393
Pages	435-47
Authors	Clark NE, Garman SC
Title	The 1.9 a structure of human alpha-N-acetylgalactosaminidase: The molecular basis of Schindler and Kanzaki diseases.
Related PDB	3h53 3h54 3h55 3igu
Related UniProtKB	P17050

Comments
This enzyme belongs to glycosidase family-27, with a retaining mechanism. For this enzyme, both substrate and product are alpha-anomers with an axial configuration at C1 atom, due to the retaining mechanism (see [4]). According to the literature [4] and [5], the reaction proceeds as follows: (1) Asp201 acts as a general acid to protonate the leaving N-acetyl-D-galactosaminide. This protonation might form an oxocarbenium-ion like transition-state. (2) Meanwhile, Asp140 (of 1ktb) makes a nucleophilic attack on the C1 atom of N-acetyl-D-galactosaminide, forming a covalent intermediate with an inverted configuration at C1. (3) Asp201 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, releasing the N-acetyl-D-galactosamine product. (X) Considering the conservation and relative location of active-site residues, Arg197 modulates the activity of Asp201, whilst Tyr103 modulates the activity of Asp140. Asp61 may stabilize the transition-state through the hydrogen bond with O4 atom.

Comments

This enzyme belongs to glycosidase family-27, with a retaining mechanism.
For this enzyme, both substrate and product are alpha-anomers with an axial configuration at C1 atom, due to the retaining mechanism (see [4]).
According to the literature [4] and [5], the reaction proceeds as follows:
(1) Asp201 acts as a general acid to protonate the leaving N-acetyl-D-galactosaminide. This protonation might form an oxocarbenium-ion like transition-state.
(2) Meanwhile, Asp140 (of 1ktb) makes a nucleophilic attack on the C1 atom of N-acetyl-D-galactosaminide, forming a covalent intermediate with an inverted configuration at C1.
(3) Asp201 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, releasing the N-acetyl-D-galactosamine product.
(X) Considering the conservation and relative location of active-site residues, Arg197 modulates the activity of Asp201, whilst Tyr103 modulates the activity of Asp140. Asp61 may stabilize the transition-state through the hydrogen bond with O4 atom.

Created	Updated
2010-03-10	2012-02-01