DB code: M00048

RLCP classification	3.100.305000.501 : Transfer
RLCP classification	1.15.8230.361 : Hydrolysis
CATH domain	3.30.565.10 : Heat Shock Protein 90	Catalytic domain
	3.30.230.10 : Ribosomal Protein S5; domain 2	Catalytic domain
	3.30.1490.30 : Dna Ligase; domain 1
	3.40.50.670 : Rossmann fold	Catalytic domain
	1.10.268.10 : Topoisomerase; domain 3
	3.30.1360.40 : Gyrase A; domain 2
	3.90.199.10 : Topoisomerase II; domain 5	Catalytic domain
E.C.	5.99.1.3
CSA
M-CSA
MACiE

CATH domain	Related DB codes (homologues)
1.10.268.10 : Topoisomerase; domain 3	M00137
3.30.1360.40 : Gyrase A; domain 2	M00137
3.30.230.10 : Ribosomal Protein S5; domain 2	T00244 M00213
3.30.565.10 : Heat Shock Protein 90	M00213
3.90.199.10 : Topoisomerase II; domain 5	M00137

Uniprot Enzyme Name
UniprotKB	Protein name	Synonyms	RefSeq	Pfam
P06786	DNA topoisomerase 2	EC 5.99.1.3 DNA topoisomerase II	NP_014311.3 (Protein) NM_001182926.3 (DNA/RNA sequence)	PF00204 (DNA_gyraseB) PF00521 (DNA_topoisoIV) PF02518 (HATPase_c) PF01751 (Toprim) [Graphical View]

KEGG enzyme name
DNA topoisomerase (ATP-hydrolysing) type II DNA topoisomerase DNA-gyrase deoxyribonucleate topoisomerase deoxyribonucleic topoisomerase topoisomerase DNA topoisomerase II

UniprotKB: Accession Number	Entry name	Activity	Subunit	Subcellular location	Cofactor
P06786	TOP2_YEAST	ATP-dependent breakage, passage and rejoining of double-stranded DNA.	Homodimer.	Nucleus.

KEGG Pathways
Map code	Pathways	E.C.

Compound table
	Cofactors	Substrates			Products			Intermediates
KEGG-id	C00305	C00434	C00002	C00001	C00434	C00008	C00009
E.C.
Compound	Magnesium	Double-stranded DNA	ATP	H2O	Double-stranded DNA	ADP	Orthophosphate
Type	divalent metal (Ca2+, Mg2+)	nucleic acids	amine group,nucleotide	H2O	nucleic acids	amine group,nucleotide	phosphate group/phosphate ion
ChEBI	18420 18420		15422 15422	15377 15377		16761 16761	26078 26078
PubChem	888 888		5957 5957	22247451 962 22247451 962		6022 6022	1004 22486802 1004 22486802

1pvgA01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1pvgB01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1q1dA01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1q1dB01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1qzrA01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1qzrB01	Bound:_MG	Unbound	Analogue:ANP		Unbound	Unbound	Unbound
1pvgA02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1pvgB02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1q1dA02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1q1dB02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1qzrA02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1qzrB02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bgwA01	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bjtA01	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bgwA02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bjtA02	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bgwA03	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bjtA03	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bgwA04	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bjtA04	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bgwA05	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound
1bjtA05	Unbound	Unbound	Unbound		Unbound	Unbound	Unbound

Reference for Active-site residues
resource	references	E.C.
Swis-prot;P06786 & literature [15] & [22]

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

1pvgA01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1pvgB01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1q1dA01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1q1dB01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1qzrA01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1qzrB01	GLU 66	ASN 70(magnesium binding)		ARG 141;ASN 142;GLY 143;TYR 144;GLY 145
1pvgA02	GLN 365;LYS 367
1pvgB02	GLN 365;LYS 367
1q1dA02	GLN 365;LYS 367
1q1dB02	GLN 365;LYS 367
1qzrA02	GLN 365;LYS 367
1qzrB02	GLN 365;LYS 367
1bgwA01
1bjtA01
1bgwA02	GLU 450;ASP 531	GLU 450;ASP 527;ASP 529;ASP 531(magnesium ion)			invisible 634-682
1bjtA02	GLU 449;	GLU 449;ASP 526; ; (magnesium ion)			invisible 527-531, 632-652, 660-674
1bgwA03
1bjtA03
1bgwA04
1bjtA04
1bgwA05	ARG 691;ARG 782;TYR 783				invisible TYR 1085;SER 1086(phospholylation)
1bjtA05	ARG 690;ARG 781;TYR 782				invisible TYR 1086;SER 1087(phospholylation)

References for Catalytic Mechanism
References	Sections	No. of steps in catalysis
[1]	Fig.1, p.271-272
[3]	Fig.3
[5]	Fig.4, p.85-87
[6]	Fig.5
[10]	Fig.3, p.157-158
[11]	Fig.2, p.142-144
[15]	p.323-325
[17]	p.886
[20]	p.271-272
[22]	p.10631

References
[1]
Resource
Comments
Medline ID
PubMed ID	1654050
Journal	Bioessays
Year	1991
Volume	13
Pages	269-73
Authors	Osheroff N, Zechiedrich EL, Gale KC
Title	Catalytic function of DNA topoisomerase II.
Related PDB
Related UniProtKB
[2]
Resource
Comments	REVIEW ON PHOSPHORYLATION.
Medline ID	93073815
PubMed ID	1332607
Journal	Antonie Van Leeuwenhoek
Year	1992
Volume	62
Pages	15-24
Authors	Gasser SM, Walter R, Dang Q, Cardenas ME
Title	Topoisomerase II: its functions and phosphorylation.
Related PDB
Related UniProtKB	P06786
[3]
Resource
Comments
Medline ID
PubMed ID	7980433
Journal	Biochem J
Year	1994
Volume	303
Pages	681-95
Authors	Watt PM, Hickson ID
Title	Structure and function of type II DNA topoisomerases.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID	7893659
Journal	Biochemistry
Year	1995
Volume	34
Pages	3632-9
Authors	Lamhasni S, Larsen AK, Barray M, Monnot M, Delain E, Fermandjian S
Title	Changes of self-association, secondary structure, and biological activity properties of topoisomerase II under varying salt conditions.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID	8696977
Journal	Curr Opin Struct Biol
Year	1996
Volume	6
Pages	84-90
Authors	Berger JM, Wang JC
Title	Recent developments in DNA topoisomerase II structure and mechanism.
Related PDB
Related UniProtKB
[6]
Resource
Comments	X-RAY CRYSTALLOGRAPHY (2.7 ANGSTROMS) OF 410-1202.
Medline ID	96138378
PubMed ID	8538787
Journal	Nature
Year	1996
Volume	379
Pages	225-32
Authors	Berger JM, Gamblin SJ, Harrison SC, Wang JC
Title	Structure and mechanism of DNA topoisomerase II.
Related PDB	1bgw
Related UniProtKB	P06786
[7]
Resource
Comments
Medline ID
PubMed ID	8610153
Journal	Proc Natl Acad Sci U S A
Year	1996
Volume	93
Pages	2975-80
Authors	Lindsley JE
Title	Intradimerically tethered DNA topoisomerase II is catalytically active in DNA transport.
Related PDB
Related UniProtKB
[8]
Resource
Comments
Medline ID
PubMed ID	8805525
Journal	Structure
Year	1996
Volume	4
Pages	117-20
Authors	Wigley DB
Title	A wasp head with a relaxing bite.
Related PDB
Related UniProtKB
[9]
Resource
Comments
Medline ID
PubMed ID	9388273
Journal	J Biol Chem
Year	1997
Volume	272
Pages	31190-5
Authors	Li W, Wang JC
Title	Footprinting of yeast DNA topoisomerase II lysyl side chains involved in substrate binding and interdomainal interactions.
Related PDB
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID	9748552
Journal	Biochim Biophys Acta
Year	1998
Volume	1400
Pages	155-71
Authors	Andoh T, Ishida R
Title	Catalytic inhibitors of DNA topoisomerase II.
Related PDB
Related UniProtKB
[11]
Resource
Comments
Medline ID
PubMed ID	9748545
Journal	Biochim Biophys Acta
Year	1998
Volume	1400
Pages	139-54
Authors	Burden DA, Osheroff N
Title	Mechanism of action of eukaryotic topoisomerase II and drugs targeted to the enzyme.
Related PDB
Related UniProtKB
[12]
Resource
Comments
Medline ID
PubMed ID	9710672
Journal	Clin Infect Dis
Year	1998
Volume	27 Suppl 1
Pages	S54-63
Authors	Hooper DC
Title	Bacterial topoisomerases, anti-topoisomerases, and anti-topoisomerase resistance.
Related PDB
Related UniProtKB
[13]
Resource
Comments
Medline ID
PubMed ID	9657678
Journal	Biochemistry
Year	1998
Volume	37
Pages	9658-67
Authors	Smith CV, Maxwell A
Title	Identification of a residue involved in transition-state stabilization in the ATPase reaction of DNA gyrase.
Related PDB
Related UniProtKB
[14]
Resource
Comments
Medline ID
PubMed ID	10419479
Journal	J Biol Chem
Year	1999
Volume	274
Pages	21688-94
Authors	Olland S, Wang JC
Title	Catalysis of ATP hydrolysis by two NH(2)-terminal fragments of yeast DNA topoisomerase II.
Related PDB
Related UniProtKB
[15]
Resource
Comments	X-ray crystallography
Medline ID
PubMed ID	10201398
Journal	Nat Struct Biol
Year	1999
Volume	6
Pages	322-6
Authors	Fass D, Bogden CE, Berger JM
Title	Quaternary changes in topoisomerase II may direct orthogonal movement of two DNA strands.
Related PDB	1bjt
Related UniProtKB
[16]
Resource
Comments
Medline ID
PubMed ID	10380229
Journal	Pac Symp Biocomput
Year	1999
Volume
Pages	578-89
Authors	Shaiu WL, Hu T, Hsieh TS
Title	The hydrophilic, protease-sensitive terminal domains of eucaryotic DNA topoisomerases have essential intracellular functions.
Related PDB
Related UniProtKB
[17]
Resource
Comments
Medline ID
PubMed ID	9927662
Journal	Proc Natl Acad Sci U S A
Year	1999
Volume	96
Pages	881-6
Authors	Liu Q, Wang JC
Title	Similarity in the catalysis of DNA breakage and rejoining by type IA and IIA DNA topoisomerases.
Related PDB
Related UniProtKB
[18]
Resource
Comments
Medline ID
PubMed ID	10713116
Journal	J Biol Chem
Year	2000
Volume	275
Pages	7980-7
Authors	Dong J, Walker J, Nitiss JL
Title	A mutation in yeast topoisomerase II that confers hypersensitivity to multiple classes of topoisomerase II poisons.
Related PDB
Related UniProtKB
[19]
Resource
Comments
Medline ID
PubMed ID	11090281
Journal	J Mol Biol
Year	2000
Volume	304
Pages	385-95
Authors	Mizushina Y, Sugawara F, Iida A, Sakaguchi K
Title	Structural homology between DNA binding sites of DNA polymerase beta and DNA topoisomerase II.
Related PDB
Related UniProtKB
[20]
Resource
Comments
Medline ID
PubMed ID	10637609
Journal	Trends Biochem Sci
Year	2000
Volume	25
Pages	24-8
Authors	Dutta R, Inouye M
Title	GHKL, an emergent ATPase/kinase superfamily.
Related PDB
Related UniProtKB
[21]
Resource
Comments
Medline ID
PubMed ID	12596227
Journal	Bioessays
Year	2003
Volume	25
Pages	232-42
Authors	Gadelle D, Filee J, Buhler C, Forterre P
Title	Phylogenomics of type II DNA topoisomerases.
Related PDB
Related UniProtKB
[22]
Resource
Comments
Medline ID
PubMed ID	12963818
Journal	Proc Natl Acad Sci U S A
Year	2003
Volume	100
Pages	10629-34
Authors	Classen S, Olland S, Berger JM
Title	Structure of the topoisomerase II ATPase region and its mechanism of inhibition by the chemotherapeutic agent ICRF-187.
Related PDB	1pvg 1q1d 1qzr
Related UniProtKB

Comments
This enzyme is composed of ATPase region, which is responsible for hydrolysis of ATP, and topoisomerase region, which is responsible for intramolecular transfer reaction of DNA molecule (DNA binding and clevage). The PDB structures, 1pvg, 1q1d, & 1qzr, correspond to the ATPase region, whilst the rest of PDB structures, 1bgw & 1bjt, correspond to the topoisomeraes region. The topoisomerase region can be divided into the N-terminal B' fragment (residues 410-679) and C-terminal A' fragment (residues 680-1202). The structure of the C-terminal domain with phosphorylation sites has not been determined yet. The literature [22] describes the roles and the relationships between the two functional regions, ATPase and topisomerase regions. According to the literature [10] & [11], the catalytic cycle of this enzyme proceeds as follows: (A) DNA binding: A pair of DNA duplexes bind to the enzyme. Here, the DNA duplex that will be cleaved is called G-segment of DNA, whilst the other DNA duplex that will be transported is called T-segment. (B) The cleavage of G-segment (pre-strand passage): At this step, active-site tyrosine (Tyr783 of 1bgw) residues at the dimer of A' fragment make nucleophilic attacks on the 5'-terminal phosphate groups of the DNA broken sites, to form covalent bonds with the G-segment, by leaving 4-base stagger. This reaction requires magnesium ion, which must be bound to cluster of acidic residues (Glu450, Asp527, Asp529 & Asp531 of 1bgw) (see [15]). According to the literature [17], this trans-esterfication proceeds as follows: (B1) Either Glu450 or Asp531 (of 1bgw) from other protomer (of the dimer) assists the active-site tyrosine, Tyr783, probably by abstracting a proton from the phenol group. (B2) The activated Tyr783 makes a nucleophilic attack on the phosphate of G-segment, to form a covalent bond. At this step, Arg690 and Arg650 (from the other chain) probably stabilize the transtion-state together with magnesium ion bound to the acidic residues (However, the tertiary structure of the dimer complex is not available, and it is difficult to confirm that these residues are close enough.) (C) DNA strand passage: ATP binding to the N-terminal ATPase domain converts the enzyme to a closed protein clamp, by inducing the dimerization of the N-terminal domain (see [5]). During the dimerization, the enzyme domains capture the second DNA duplex (T-segment), and push it into the interior of the enzyme and through the DNA gate, made by the cleavage of the first DNA duplex (G-segment) (see [6]). (B') The religation of G-segment (post-strand passage): The reverse reaction of the cleavage. (Possibly, the hydroxyl group of 3'-end of DNA makes a nucleophilic attack on the phosphate group linked to the active-site tyrosine residues.) (D) ATP hydrolysis: ATP hydrolysis triggers the opening of the protein clamp, releasing the DNA product (T-segment). According to the literature [13] & [20], the ATP hydrolysis (at ATPase) proceeds as follows; (D1) Glu66 (of 1pvg) acts as a general base, to activate a water molecule, which must be in-line for a nucleophilic attack on the gamma-phosphate of ATP. Lys367 may stabilize the activated water during this reaction. (D2) Lys367 stabilizes the transition-state, along with mainchain amide of resdiues 141-145. Magnesium ion, bound to Asn70, stabilizes the transition-state.

Comments

This enzyme is composed of ATPase region, which is responsible for hydrolysis of ATP, and topoisomerase region, which is responsible for intramolecular transfer reaction of DNA molecule (DNA binding and clevage). The PDB structures, 1pvg, 1q1d, & 1qzr, correspond to the ATPase region, whilst the rest of PDB structures, 1bgw & 1bjt, correspond to the topoisomeraes region. The topoisomerase region can be divided into the N-terminal B' fragment (residues 410-679) and C-terminal A' fragment (residues 680-1202). The structure of the C-terminal domain with phosphorylation sites has not been determined yet.
The literature [22] describes the roles and the relationships between the two functional regions, ATPase and topisomerase regions.
According to the literature [10] & [11], the catalytic cycle of this enzyme proceeds as follows:
(A) DNA binding: A pair of DNA duplexes bind to the enzyme. Here, the DNA duplex that will be cleaved is called G-segment of DNA, whilst the other DNA duplex that will be transported is called T-segment.
(B) The cleavage of G-segment (pre-strand passage): At this step, active-site tyrosine (Tyr783 of 1bgw) residues at the dimer of A' fragment make nucleophilic attacks on the 5'-terminal phosphate groups of the DNA broken sites, to form covalent bonds with the G-segment, by leaving 4-base stagger. This reaction requires magnesium ion, which must be bound to cluster of acidic residues (Glu450, Asp527, Asp529 & Asp531 of 1bgw) (see [15]).
According to the literature [17], this trans-esterfication proceeds as follows:
(B1) Either Glu450 or Asp531 (of 1bgw) from other protomer (of the dimer) assists the active-site tyrosine, Tyr783, probably by abstracting a proton from the phenol group.
(B2) The activated Tyr783 makes a nucleophilic attack on the phosphate of G-segment, to form a covalent bond. At this step, Arg690 and Arg650 (from the other chain) probably stabilize the transtion-state together with magnesium ion bound to the acidic residues
(However, the tertiary structure of the dimer complex is not available, and it is difficult to confirm that these residues are close enough.)
(C) DNA strand passage: ATP binding to the N-terminal ATPase domain converts the enzyme to a closed protein clamp, by inducing the dimerization of the N-terminal domain (see [5]). During the dimerization, the enzyme domains capture the second DNA duplex (T-segment), and push it into the interior of the enzyme and through the DNA gate, made by the cleavage of the first DNA duplex (G-segment) (see [6]).
(B') The religation of G-segment (post-strand passage): The reverse reaction of the cleavage. (Possibly, the hydroxyl group of 3'-end of DNA makes a nucleophilic attack on the phosphate group linked to the active-site tyrosine residues.)
(D) ATP hydrolysis: ATP hydrolysis triggers the opening of the protein clamp, releasing the DNA product (T-segment). According to the literature [13] & [20], the ATP hydrolysis (at ATPase) proceeds as follows;
(D1) Glu66 (of 1pvg) acts as a general base, to activate a water molecule, which must be in-line for a nucleophilic attack on the gamma-phosphate of ATP. Lys367 may stabilize the activated water during this reaction.
(D2) Lys367 stabilizes the transition-state, along with mainchain amide of resdiues 141-145. Magnesium ion, bound to Asn70, stabilizes the transition-state.

Created	Updated
2004-04-25	2010-02-08