DB code: S00510

RLCP classification 2.40.18000.65 : Phosphorolysis
CATH domain 3.40.50.1580 : Rossmann fold Catalytic domain
E.C. 2.4.2.1
CSA 1a69
M-CSA 1a69
MACiE

CATH domain Related DB codes (homologues)
3.40.50.1580 : Rossmann fold S00375 S00376

Uniprot Enzyme Name
UniprotKB Protein name Synonyms RefSeq Pfam
P0ABP8 Purine nucleoside phosphorylase deoD-type
PNP
EC 2.4.2.1
Inosine phosphorylase
NP_418801.1 (Protein)
NC_000913.2 (DNA/RNA sequence)
YP_492514.1 (Protein)
NC_007779.1 (DNA/RNA sequence)
PF01048 (PNP_UDP_1)
[Graphical View]

KEGG enzyme name
purine-nucleoside phosphorylase
inosine phosphorylase
PNPase
PUNPI
PUNPII
inosine-guanosine phosphorylase
nucleotide phosphatase
purine deoxynucleoside phosphorylase
purine deoxyribonucleoside phosphorylase
purine nucleoside phosphorylase
purine ribonucleoside phosphorylase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P0ABP8 DEOD_ECOLI Purine nucleoside + phosphate = purine + alpha-D-ribose 1-phosphate. Homohexamer.

KEGG Pathways
Map code Pathways E.C.
MAP00230 Purine metabolism
MAP00240 Pyrimidine metabolism
MAP00760 Nicotinate and nicotinamide metabolism

Compound table
Substrates Products Intermediates
KEGG-id C15586 C00009 C15587 C00620
E.C.
Compound Purine nucleoside Orthophosphate Purine alpha-D-Ribose 1-phosphate
Type nucleoside phosphate group/phosphate ion aromatic ring (with nitrogen atoms) carbohydrate,phosphate group/phosphate ion
ChEBI 18255
26078
17258
35586
35588
35589
16300
PubChem 68368
1004
22486802
1044
439236
1a69A Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FMB Analogue:SO4 Unbound Unbound
1a69B Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FMB Analogue:SO4 Unbound Unbound
1a69C Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FMB Analogue:SO4 Unbound Unbound
1ecpA Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1ecpB Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1ecpC Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1ecpD Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1ecpE Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1ecpF Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1k9sA Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM2 Bound:PO4 Unbound Unbound
1k9sB Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM2 Bound:PO4 Unbound Unbound
1k9sC Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM2 Bound:PO4 Unbound Unbound
1k9sD Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM1 Bound:PO4 Unbound Unbound
1k9sE Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM1 Bound:PO4 Unbound Unbound
1k9sF Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:FM1 Bound:PO4 Unbound Unbound

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

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1a69A Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1a69B Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1a69C Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpA Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpB Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpC Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpD Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpE Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpF Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sA Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sB Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sC Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sD Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sE Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sF Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain ASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[1]
p.1380
[3]
Fig.1, Fig.8 2
[5]
Scheme 6, p.363-365 2
[6]
Fig.5, Fig.7, p.14 2

References
[1]
Resource
Comments
Medline ID
PubMed ID 9351810
Journal Structure
Year 1997
Volume 5
Pages 1373-83
Authors Mao C, Cook WJ, Zhou M, Koszalka GW, Krenitsky TA, Ealick SE
Title The crystal structure of Escherichia coli purine nucleoside phosphorylase: a comparison with the human enzyme reveals a conserved topology.
Related PDB 1ecp
Related UniProtKB P0ABP8
[2]
Resource
Comments
Medline ID
PubMed ID 9653038
Journal J Mol Biol
Year 1998
Volume 280
Pages 153-66
Authors Koellner G, Luic M, Shugar D, Saenger W, Bzowska A
Title Crystal structure of the ternary complex of E. coli purine nucleoside phosphorylase with formycin B, a structural analogue of the substrate inosine, and phosphate (Sulphate) at 2.1 A resolution.
Related PDB 1a69
Related UniProtKB P0ABP8
[3]
Resource
Comments
Medline ID
PubMed ID 11444966
Journal Biochemistry
Year 2001
Volume 40
Pages 8204-15
Authors Shi W, Basso LA, Santos DS, Tyler PC, Furneaux RH, Blanchard JS, Almo SC, Schramm VL
Title Structures of purine nucleoside phosphorylase from Mycobacterium tuberculosis in complexes with immucillin-H and its pieces.
Related PDB
Related UniProtKB
[4]
Resource
Comments
Medline ID
PubMed ID 11591349
Journal Structure (Camb)
Year 2001
Volume 9
Pages 941-53
Authors Lee JE, Cornell KA, Riscoe MK, Howell PL
Title Structure of E. coli 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase reveals similarity to the purine nucleoside phosphorylases.
Related PDB
Related UniProtKB
[5]
Resource
Comments
Medline ID
PubMed ID 11786017
Journal J Mol Biol
Year 2002
Volume 315
Pages 351-71
Authors Koellner G, Bzowska A, Wielgus-Kutrowska B, Luic M, Steiner T, Saenger W, Stepinski J
Title Open and closed conformation of the E. coli purine nucleoside phosphorylase active center and implications for the catalytic mechanism.
Related PDB
Related UniProtKB
[6]
Resource
Comments
Medline ID
PubMed ID 11743878
Journal Biochem J
Year 2002
Volume 361
Pages 1-25
Authors Pugmire MJ, Ealick SE
Title Structural analyses reveal two distinct families of nucleoside phosphorylases.
Related PDB
Related UniProtKB

Comments
The enzymes of this entry corresponds to the hexameric subunit members of nucleoside phosphorylase family-I [6].
FMB molecules (analogues of purine nucleoside) in 1a69 interact with two chains (A & C).
The literature [6] summarized the proposed catalytic mechanism of the enzyme. Beta-nucleoside binds in a high-energy (anticlinal torsion angle of the glycosidic bond, with the ribose moiety in the uncommon C-4'-endo sugar pucker), according to the literature. This high-energy conformation produces steric strain, which induces glycosidic cleavage. The glycosidic bond is weakened further as electrons flow from O-4' of the ribose to the purine ring, resulting in an oxocarbenium ion that is stabilized by the negative charges of the phosphate ion. The phosphate ion binds on the alpha-side of the ribose ring, where it is postioned to participate in an SN1 nucleophilic attack at the C-1' position. The flow of electrons from the glycosidic bond to the purine ring is probably stabilized by active site residues (Asp) interactions at O-4 position of the purine base.
According to the literature [5], the hexamer of this enzymes is found as a trimer of unsymmetric dimers, which are formed by pairs of monomers with active sites in different conformations. The conformational differences results from a flexible helix (H8: residues 214-236), which is continuous in one conformer, and segmented in the other. With the continuous helix, the entry in to the active site pocket is wide open, and the ligands are bound only loosely ("open" conformation). In contrast, by segmentation of the helix (H8: residues 214-219 and H8': residues 223-236, separated), the entry in to the active site is partially closed, the pocket is narrowed and the ligands are bound much more tightly ("closed" conformation) [5].
The possible catalytic mechanism could be described as follows [5]:
(1) In the open conformation, the helix H8 is continuous and Arg217 on the helix is far away from Asp204 which can be in the acid form. The sidechain of Arg24 is floppy. Phosphate and the nucleoside both can bind to this conformation of the active site. The neutral sidechain of Asp204 donates a hydrogen bond to the purine nitrogen N7.
(2) Phosphate binding stabilizes Arg24 and, in turn, favors breaking of H8 into two segments. The active site is now in the closed conformation.
(3) The conformation change brings the guanidinium group of Arg217 in contact with the acid group of Asp204. This triggers proton transfer to the base, followed by formation of a salt bridge between Arg217 and Asp204. The positively charged purine base leads to the ribo-oxocarbenium ion character of the ribose, representing the transition state of bond cleavage in acid hydorolysis which can then proceed further.

Created Updated
2002-07-12 2011-09-27