EzCatDB: S00510
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DB codeS00510
RLCP classification2.40.18000.65 : Phosphorolysis
CATH domainDomain 13.40.50.1580 : Rossmann foldCatalytic domain
E.C.2.4.2.1
CSA1a69

CATH domainRelated DB codes (homologues)
3.40.50.1580 : Rossmann foldS00375,S00376

Enzyme Name
UniProtKBKEGG

P0ABP8
Protein namePurine nucleoside phosphorylase deoD-typepurine-nucleoside phosphorylase
inosine phosphorylase
PNPase
PUNPI
PUNPII
inosine-guanosine phosphorylase
nucleotide phosphatase
purine deoxynucleoside phosphorylase
purine deoxyribonucleoside phosphorylase
purine nucleoside phosphorylase
purine ribonucleoside phosphorylase
SynonymsPNP
EC 2.4.2.1
Inosine phosphorylase
RefSeqNP_418801.1 (Protein)
NC_000913.2 (DNA/RNA sequence)
YP_492514.1 (Protein)
NC_007779.1 (DNA/RNA sequence)
PfamPF01048 (PNP_UDP_1)
[Graphical view]

KEGG pathways
MAP codePathways
MAP00230Purine metabolism
MAP00240Pyrimidine metabolism
MAP00760Nicotinate and nicotinamide metabolism

UniProtKB:Accession NumberP0ABP8
Entry nameDEOD_ECOLI
ActivityPurine nucleoside + phosphate = purine + alpha-D-ribose 1-phosphate.
SubunitHomohexamer.
Subcellular location
Cofactor

Compound table: links to PDB-related databases & PoSSuM

SubstratesProducts
KEGG-idC15586C00009C15587C00620
CompoundPurine nucleosideOrthophosphatePurinealpha-D-Ribose 1-phosphate
Typenucleosidephosphate group/phosphate ionaromatic ring (with nitrogen atoms)carbohydrate,phosphate group/phosphate ion
ChEBI18255
26078
35589
35588
35586
17258
16300
PubChem68368
22486802
1004
1044
439236
            
1a69AAnalogue:FMBAnalogue:SO4UnboundUnbound
1a69BAnalogue:FMBAnalogue:SO4UnboundUnbound
1a69CAnalogue:FMBAnalogue:SO4UnboundUnbound
1ecpAUnboundUnboundUnboundUnbound
1ecpBUnboundUnboundUnboundUnbound
1ecpCUnboundUnboundUnboundUnbound
1ecpDUnboundUnboundUnboundUnbound
1ecpEUnboundUnboundUnboundUnbound
1ecpFUnboundUnboundUnboundUnbound
1k9sAAnalogue:FM2Bound:PO4UnboundUnbound
1k9sBAnalogue:FM2Bound:PO4UnboundUnbound
1k9sCAnalogue:FM2Bound:PO4UnboundUnbound
1k9sDAnalogue:FM1Bound:PO4UnboundUnbound
1k9sEAnalogue:FM1Bound:PO4UnboundUnbound
1k9sFAnalogue:FM1Bound:PO4UnboundUnbound

Active-site residues
resource
literature [5]
pdbCatalytic residues
         
1a69AASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1a69BASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1a69CASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpAASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpBASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpCASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpDASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpEASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1ecpFASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sAASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sBASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sCASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sDASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sEASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)
1k9sFASP 204(base binding);ARG 24(phosphate binding);ARG 217(Switch)

References for Catalytic Mechanism
ReferencesSectionsNo. of steps in catalysis
[1]p.1380
[3]Fig.1, Fig.82
[5]Scheme 6, p.363-3652
[6]Fig.5, Fig.7, p.142

references
[1]
PubMed ID9351810
JournalStructure
Year1997
Volume5
Pages1373-83
AuthorsMao C, Cook WJ, Zhou M, Koszalka GW, Krenitsky TA, Ealick SE
TitleThe crystal structure of Escherichia coli purine nucleoside phosphorylase: a comparison with the human enzyme reveals a conserved topology.
Related PDB1ecp
Related UniProtKBP0ABP8
[2]
PubMed ID9653038
JournalJ Mol Biol
Year1998
Volume280
Pages153-66
AuthorsKoellner G, Luic M, Shugar D, Saenger W, Bzowska A
TitleCrystal 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 PDB1a69
Related UniProtKBP0ABP8
[3]
PubMed ID11444966
JournalBiochemistry
Year2001
Volume40
Pages8204-15
AuthorsShi W, Basso LA, Santos DS, Tyler PC, Furneaux RH, Blanchard JS, Almo SC, Schramm VL
TitleStructures of purine nucleoside phosphorylase from Mycobacterium tuberculosis in complexes with immucillin-H and its pieces.
[4]
PubMed ID11591349
JournalStructure (Camb)
Year2001
Volume9
Pages941-53
AuthorsLee JE, Cornell KA, Riscoe MK, Howell PL
TitleStructure of E. coli 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase reveals similarity to the purine nucleoside phosphorylases.
[5]
PubMed ID11786017
JournalJ Mol Biol
Year2002
Volume315
Pages351-71
AuthorsKoellner G, Bzowska A, Wielgus-Kutrowska B, Luic M, Steiner T, Saenger W, Stepinski J
TitleOpen and closed conformation of the E. coli purine nucleoside phosphorylase active center and implications for the catalytic mechanism.
[6]
PubMed ID11743878
JournalBiochem J
Year2002
Volume361
Pages1-25
AuthorsPugmire MJ, Ealick SE
TitleStructural analyses reveal two distinct families of nucleoside phosphorylases.

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.

createdupdated
2002-07-122011-09-27


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