DB code: S00214

RLCP classification 1.60.55100.9000 : Hydrolysis
CATH domain 3.20.20.80 : TIM Barrel Catalytic domain
E.C. 3.2.1.147
CSA 1myr
M-CSA 1myr
MACiE

CATH domain Related DB codes (homologues)
3.20.20.80 : TIM Barrel S00202 S00210 S00748 S00906 S00907 S00911 S00912 S00915 M00134 M00160 D00479 S00204 S00205 S00206 S00207 S00203 S00208 S00209 S00211 S00213 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 CAZy Pfam
P29736 Myrosinase MA1
EC 3.2.1.147
Sinigrinase
Thioglucosidase
GH1 (Glycoside Hydrolase Family 1)
PF00232 (Glyco_hydro_1)
[Graphical View]

KEGG enzyme name
thioglucosidase
myrosinase
sinigrinase
sinigrase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P29736 MYRA_SINAL A thioglucoside + H(2)O = a sugar + a thiol. Homodimer. Vacuole. Binds 1 ascorbate molecule per subunit.

KEGG Pathways
Map code Pathways E.C.
MAP00380 Tryptophan metabolism

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00072 C02085 C00001 C00145 C11477
E.C.
Compound Ascorbate Thioglucoside H2O Thiol Sugar Transition-state in glycosylation Intermediate Transition-state in deglycosylation
Type carbohydrate,aromatic ring (with hetero atoms other than nitrogen atoms) carbohydrate,sulfide group H2O sulfhydryl group carbohydrate
ChEBI 29073
 29073
 		15377
 15377
PubChem 54670067
 54670067
 		22247451
 962
 22247451
 962
1dwaM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1dwfM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1dwgM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1dwhM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1dwiM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1dwjM Unbound Unbound Unbound Analogue:GOL_24 Unbound Unbound Unbound
1e4mM Unbound Unbound Unbound Analogue:GOL_1515 Unbound Unbound Unbound
1e6qM Unbound Unbound Unbound Unbound Transition-state-analogue:NTZ Unbound Unbound
1e6sM Unbound Unbound Unbound Unbound Transition-state-analogue:GOX Unbound Unbound
1e6xM Unbound Unbound Unbound Unbound Transition-state-analogue:LGC Unbound Unbound
1e70M Unbound Unbound Unbound Unbound Unbound Unbound Transition-state-analogue:G2F
1e71M Bound:ASC Unbound Unbound Unbound Unbound Unbound Unbound
1e72M Bound:ASC Unbound Unbound Unbound Transition-state-analogue:GOX Unbound Unbound
1e73M Bound:ASC Unbound Bound:HOH_998 Unbound Unbound Unbound Intermediate-analogue:G2F Unbound
1myrA Unbound Unbound Unbound Analogue:GOL_22 Unbound Unbound Unbound
2myrA Unbound Unbound Unbound Unbound Unbound Intermediate-analogue:G2F Unbound

Reference for Active-site residues
resource references E.C.
PDB;1e6q & literature [6], [9]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1dwaM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1dwfM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1dwgM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1dwhM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1dwiM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1dwjM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e4mM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e6qM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e6sM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e6xM GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e70M GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e71M GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e72M GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1e73M GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
1myrA GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)
2myrA GLN 187;TYR 330;GLU 409 GLN 187;ARG 259(Ascorbate binding)

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[3]
Fig.3, p.15257
[4]
Fig.4, p89
[6]
Fig.1, p.671-673
[9]
FIG.8, p39388-39393
[10]
Fig.8, p.648

References
[1]
Resource
Comments
Medline ID
PubMed ID 7759287
Journal Hereditas
Year 1995
Volume 122
Pages 95-8
Authors Taipalensuu J, Lundgren S, Rask L
Title No evidence for the ascorbic activation site of myrosinase being encoded by end of exon 9, beginning of exon 10.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID 7657629
Journal J Biol Chem
Year 1995
Volume 270
Pages 20530-5
Authors Botti MG, Taylor MG, Botting NP
Title Studies on the mechanism of myrosinase. Investigation of the effect of glycosyl acceptors on enzyme activity.
Related PDB
Related UniProtKB
[3]
Resource
Comments
Medline ID
PubMed ID 8952475
Journal Biochemistry
Year 1996
Volume 35
Pages 15256-9
Authors Cottaz S, Henrissat B, Driguez H
Title Mechanism-based inhibition and stereochemistry of glucosinolate hydrolysis by myrosinase.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 8641474
Journal FEBS Lett
Year 1996
Volume 385
Pages 87-90
Authors Iori R, Rollin P, Streicher H, Thiem J, Palmieri S
Title The myrosinase-glucosinolate interaction mechanism studied using some synthetic competitive inhibitors.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 9354235
Journal Bioorg Med Chem
Year 1997
Volume 5
Pages 1799-806
Authors Leoni O, Iori R, Palmieri S, Esposito E, Menegatti E, Cortesi R, Nastruzzi C
Title Myrosinase-generated isothiocyanate from glucosinolates: isolation, characterization and in vitro antiproliferative studies.
Related PDB
Related UniProtKB
[6]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.64 ANGSTROMS).
Medline ID
PubMed ID 9195886
Journal Structure
Year 1997
Volume 5
Pages 663-75
Authors Burmeister WP, Cottaz S, Driguez H, Iori R, Palmieri S, Henrissat B
Title The crystal structures of Sinapis alba myrosinase and a covalent glycosyl-enzyme intermediate provide insights into the substrate recognition and active-site machinery of an S-glycosidase.
Related PDB 1myr 2myr
Related UniProtKB Q7SIB0
[7]
Resource
Comments
Medline ID
PubMed ID 10417337
Journal Biochem J
Year 1999
Volume 341
Pages 725-32
Authors Shikita M, Fahey JW, Golden TR, Holtzclaw WD, Talalay P
Title An unusual case of 'uncompetitive activation' by ascorbic acid: purification and kinetic properties of a myrosinase from Raphanus sativus seedlings.
Related PDB
Related UniProtKB
[8]
Resource
Comments X-ray crystallography
Medline ID
PubMed ID 10713520
Journal Acta Crystallogr D Biol Crystallogr
Year 2000
Volume 56
Pages 328-41
Authors Burmeister WP
Title Structural changes in a cryo-cooled protein crystal owing to radiation damage.
Related PDB 1dwa 1dwf 1dwg 1dwh 1dwi 1dwj
Related UniProtKB
[9]
Resource
Comments X-ray crystallography
Medline ID
PubMed ID 10978344
Journal J Biol Chem
Year 2000
Volume 275
Pages 39385-93
Authors Burmeister WP, Cottaz S, Rollin P, Vasella A, Henrissat B
Title High resolution X-ray crystallography shows that ascorbate is a cofactor for myrosinase and substitutes for the function of the catalytic base.
Related PDB 1e4m 1e6q 1e6s 1e6x 1e70 1e71 1e72 1e73
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID 11738173
Journal Curr Opin Chem Biol
Year 2001
Volume 5
Pages 643-9
Authors Zechel DL, Withers SG
Title Dissection of nucleophilic and acid-base catalysis in glycosidases.
Related PDB
Related UniProtKB
[11]
Resource
Comments
Medline ID
PubMed ID 11804799
Journal Insect Biochem Mol Biol
Year 2002
Volume 32
Pages 275-84
Authors Jones AM, Winge P, Bones AM, Cole R, Rossiter JT
Title Characterization and evolution of a myrosinase from the cabbage aphid Brevicoryne brassicae.
Related PDB
Related UniProtKB
[12]
Resource
Comments
Medline ID
PubMed ID 12696966
Journal J Agric Food Chem
Year 2003
Volume 51
Pages 2737-44
Authors Bernardi R, Finiguerra MG, Rossi AA, Palmieri S
Title Isolation and biochemical characterization of a basic myrosinase from ripe Crambe abyssinica seeds, highly specific for epi-progoitrin.
Related PDB
Related UniProtKB

Comments
This enzyme was transferred from E.C. 3.2.3.1 to E.C. 3.2.1.147.
This enzyme belongs to glycosidase family-1, a member family of 4/7 superfamily, which has got catalytic residues at the C-terminal ends of beta-4 and beta-7 on the (alpha/beta)8 barrel fold.
Moreover, comparing the structural data with that of the other family-1 enzyme, cellulase (E.C. 3.2.1.4) (S00203 in EzCatDB), Tyr330 might stabilize the leaving nucleophile, Glu409 in deglycosylation.
According to the literature [6] & [10], the S-glycoside is a good leaving group, so that a general acid is not required for the first step of Enzyme-substrate intermediate formation. On the other hand, at the second step of the hydrolysis, O3 atom of the ascorbate acts as a general base, which activates the water for the hydrolysis of the intermeidate (see [9] & [10]).
The catalytic reaction proceeds as follows (see [9] & [10]):
(1) Glu409 makes a nucleophilic attack on the C1 atom of the S-glycoside, releasing the thiolate, and forming a glycosyl-enzyme intermediate.
(2) An organic cofactor, Ascorbate, acts as a general base, to activate a water. Here, Gln187 seems to modulate the O3 oxygen of ascorbate by interacting with it.
(3) The activated water makes a nucleophilic attack on the C1 atom of the intermediate. Here, Tyr330 might stabilize the leaving Glu409.

Created Updated
2005-03-23 2009-02-26