EzCatDB: T00063

DB codeT00063
RLCP classification1.32.5000.73
CATH domainDomain 12.60.40.10
Domain 23.20.20.80Catalytic domain
Domain 33.10.50.10
E.C.3.2.1.14
CSA1ctn

CATH domainRelated DB codes (homologues)
2.60.40.10M00131,T00257,T00005,M00113,M00127,M00132,M00323,M00325,M00327,M00329,M00330,M00331,M00332,T00307,D00166,D00500,M00112,M00193,T00065,T00067,T00245
3.10.50.10M00134
3.20.20.80S00202,S00210,S00748,S00906,S00907,S00911,S00912,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,T00066,T00067

Enzyme Name
Swiss-protKEGG

P07254
Protein nameChitinase Achitinase
chitodextrinase
1,4-beta-poly-N-acetylglucosaminidase
poly-beta-glucosaminidase
beta-1,4-poly-N-acetyl glucosamidinase
poly[1,4-(N-acetyl-beta-D-glucosaminide)] glycanohydrolase
SynonymsEC 3.2.1.14

KEGG pathways
MAP codePathways
MAP00530Aminosugars metabolism

Swiss-prot:Accession NumberP07254
Entry nameCHIA_SERMA
ActivityRandom hydrolysis of N-acetyl-beta-D- glucosaminide (1->4)-beta-linkages in chitin and chitodextrins.
Subunit
Subcellular location
Cofactor


SubstratesProductsintermediates
KEGG-idC00461C00851C00001C03518C00140C00461
CompoundChitinChitodextrinH2ON-Acetyl-D-glucosaminideN-Acetyl-D-glucosamineChitin
Typeamide group,polysaccharideamide group,polysaccharideH2Oamide group,carbohydrateamide group,carbohydrateamide group,polysaccharide
1ctnA01UnboundUnbound
UnboundUnboundUnboundUnbound
1edqA01UnboundUnbound
UnboundUnboundUnboundUnbound
1ehnA01UnboundUnbound
UnboundUnboundUnboundUnbound
1eibA01UnboundUnbound
UnboundUnboundUnboundUnbound
1ffrA01UnboundUnbound
UnboundUnboundUnboundUnbound
1k9tA01UnboundUnbound
UnboundUnboundUnboundUnbound
1nh6A01UnboundUnbound
UnboundUnboundUnboundUnbound
1ctnA02UnboundUnbound
UnboundUnboundUnboundUnbound
1edqA02UnboundUnbound
UnboundUnboundUnboundUnbound
1ehnA02Bound:NAG-NAG-NAG-NAG-NAG-NAG-NAG-NAGUnbound
UnboundUnboundUnboundUnbound
1eibA02Bound:NAG-NAG-NAG-NAG-NAG-NAG-NAG-NAGUnbound
UnboundUnboundUnboundUnbound
1ffrA02UnboundUnbound
UnboundUnboundUnboundTransition-state-bound:NAG-NAG-NAG-NAG-NAG(chain B),NAG-NAG(chain C)
1k9tA02Bound:NAG-NAG-NAG-NAGUnbound
UnboundUnboundUnboundUnbound
1nh6A02Bound:NAG-NAG-NAG-NAG-NAG-NAGUnbound
UnboundUnboundUnboundUnbound
1ctnA03UnboundUnbound
UnboundUnboundUnboundUnbound
1edqA03UnboundUnbound
UnboundUnboundUnboundUnbound
1ehnA03UnboundUnbound
UnboundUnboundUnboundUnbound
1eibA03UnboundUnbound
UnboundUnboundUnboundUnbound
1ffrA03UnboundUnbound
UnboundUnboundUnboundUnbound
1k9tA03UnboundUnbound
UnboundUnboundUnboundUnbound
1nh6A03UnboundUnbound
UnboundUnboundUnboundUnbound

Active-site residues
resource
literature [6]
pdbCatalytic residuescomment
1ctnA01

1edqA01

1ehnA01

1eibA01

1ffrA01

1k9tA01

1nh6A01

1ctnA02ASP 313;GLU 315;TYR 390;ASP 391

1edqA02ASP 313;GLU 315;TYR 390;ASP 391

1ehnA02ASP 313;       ;TYR 390;ASP 391
mutant E315Q
1eibA02       ;GLU 315;TYR 390;ASP 391
mutant D313A
1ffrA02ASP 313;GLU 315;       ;ASP 391
mutant Y390F
1k9tA02ASP 313;GLU 315;TYR 390;       
mutant D391A
1nh6A02ASP 313;       ;TYR 390;ASP 391
mutant E315L
1ctnA03

1edqA03

1ehnA03

1eibA03

1ffrA03

1k9tA03

1nh6A03


References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[4]p.202-203
[6]Scheme 1, p.11341-113432
[8]p.403

references
[1]
CommentsREVISIONS, AND X-RAY CRYSTALLOGRAPHY (2.3 ANGSTROMS).
Medline ID95219379
PubMed ID7704527
JournalStructure
Year1994
Volume2
Pages1169-80
AuthorsPerrakis A, Tews I, Dauter Z, Oppenheim AB, Chet I, Wilson KS, Vorgias CE
TitleCrystal structure of a bacterial chitinase at 2.3 A resolution.
Related PDB1ctn
Related Swiss-protP07254
[2]
PubMed ID8831791
JournalJ Mol Biol
Year1996
Volume262
Pages243-57
AuthorsTerwisscha van Scheltinga AC, Hennig M, Dijkstra BW
TitleThe 1.8 A resolution structure of hevamine, a plant chitinase/lysozyme, and analysis of the conserved sequence and structure motifs of glycosyl hydrolase family 18.
[3]
PubMed ID9377712
JournalFold Des
Year1997
Volume2
Pages291-4
AuthorsPerrakis A, Ouzounis C, Wilson KS
TitleEvolution of immunoglobulin-like modules in chitinases: their structural flexibility and functional implications.
[4]
PubMed ID10794597
JournalIUBMB Life
Year1999
Volume48
Pages199-204
AuthorsLin FP, Chen HC, Lin CS
TitleSite-directed mutagenesis of Asp313, Glu315, and Asp391 residues in chitinase of Aeromonas caviae.
[5]
PubMed ID10823940
JournalProc Natl Acad Sci U S A
Year2000
Volume97
Pages5842-7
Authorsvan Aalten DM, Synstad B, Brurberg MB, Hough E, Riise BW, Eijsink VG, Wierenga RK
TitleStructure of a two-domain chitotriosidase from Serratia marcescens at 1.9-A resolution.
[6]
CommentsX-ray crystallography
PubMed ID11560481
JournalBiochemistry
Year2001
Volume40
Pages11338-43
AuthorsPapanikolau Y, Prag G, Tavlas G, Vorgias CE, Oppenheim AB, Petratos K
TitleHigh resolution structural analyses of mutant chitinase A complexes with substrates provide new insight into the mechanism of catalysis.
Related PDB1ehn,1eib,1ffr
[7]
PubMed ID11342059
JournalBiochim Biophys Acta
Year2001
Volume1545
Pages349-56
AuthorsLonhienne T, Baise E, Feller G, Bouriotis V, Gerday C
TitleEnzyme activity determination on macromolecular substrates by isothermal titration calorimetry: application to mesophilic and psychrophilic chitinases.
[8]
CommentsX-ray crystallography
PubMed ID12554965
JournalActa Crystallogr D Biol Crystallogr
Year2003
Volume59
Pages400-3
AuthorsPapanikolau Y, Tavlas G, Vorgias CE, Petratos K
TitleDe novo purification scheme and crystallization conditions yield high-resolution structures of chitinase A and its complex with the inhibitor allosamidin.
Related PDB1edq
[9]
PubMed ID12932195
JournalBiochem J
Year2003
Volume376
Pages87-95
AuthorsAronson NN Jr, Halloran BA, Alexyev MF, Amable L, Madura JD, Pasupulati L, Worth C, Van Roey P
TitleFamily 18 chitinase-oligosaccharide substrate interaction: subsite preference and anomer selectivity of Serratia marcescens chitinase A.
Related PDB1nh6

comments
This enzyme is composed of the N-terminal immunoglobulin-like region and the C-terminal catalytic region, which has also two distinct domains. According to the literature [3], the N-terminal immunoglobulin-like domain might interact with the chitin substrate during catalysis.
In the early study [4], Asp313, Glu315 and Asp391 have been thought to be involved in catalysis. The paper [4] suggested that Asp313 and Glu315 might act as a stabilizer and a general acid, respectively.
According to the more recent work ([6] & [8]), however, Tyr390 was reported to be involved in the catalytic reaction, instead of Asp391. The paper [6] proposed the following mechanism:
(1) The sugar unit at the subsite (-1) adopts a boat conformation, which has higher free energy by 8kcal/mol, and its N-acetyl group points toward Asp313 and Glu315. The side chain of Asp313 points toward Asp311, away from Glu315. A water molecule is hydrogen-bonded to the phenol hydroxyl group of Tyr390 and the amine group of the acetamido group of the sugar unit at the subsite (-1). The sidechain carboxylate of Glu315 is protonated.
(2) Glu315 functions as a general acid, and protonates the O4 oxygen of the sugar unit at the subsite (+1), thus cleaving the glycosidic linkage, O4(+1)-C1(-1). (This suggests SN1-like cleavage.)
(3) The positive charge is developed on C1 and O5 atoms at the subsite (-1), which can be stabilized by the water molecule hydrogen-bonded to Tyr390 and the acetamido group at (-1), the O7 atom of the acetamido group at (-1), and the deprotonated sidechain of Glu315.
(4) This cleavage induces the rotation of Asp313 toward Glu315, and the rotation of the acetamido group at (-1) toward Tyr390. The rotation of the acetamido group translocates the water molecule bound to Tyr390 closer to Glu315.
(5) The deprotonated sidechain of Glu315 acts as a general base, abstracting the water molecule, which can act on the C1 carbon at subsite (-1).
Taken together, Glu315 acts as an acid-base, whilst Asp313 modulates the role of Glu315. Moreover, Tyr390 stabilizes the transition-state through the water. The stabilization might be assisted by N-acetyl group of the substrate.
However, the mutation of Asp391 inactivates this enzyme, suggesting that the residue must be involved in catalysis (see Table 1 of [6]).

createdupdated
2004-04-302009-02-26


Copyright: Nozomi Nagano, JST & CBRC-AIST
Funded by PRESTO/Japan Science and Technology Corporation (JST) (December 2001 - November 2004)
Funded by Grant-in-Aid for Publication of Scientific Research Results/Japan Society for the Promotion of Science (JSPS) (April 2005 - March 2006)
Funded by Grant-in-Aid for Scientific Research (B)/Japan Society for the Promotion of Science (JSPS) (April 2005 - March 2008)
Funded by BIRD/Japan Science and Technology Corporation (JST) (September 2005 - September 2010)
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)

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