3GB9

Human purine nucleoside phosphorylase double mutant E201Q,N243D complexed with 2-fluoroadenine

Summary for 3GB9

• Entry DOI: 10.2210/pdb3gb9/pdb
• Descriptor: Purine nucleoside phosphorylase, 2-fluoroadenine, SULFATE ION, ... (5 entities in total)
• Functional Keywords: enzyme-product complex, disease mutation, glycosyltransferase, transferase
• Biological source: Homo sapiens (Human)
• Cellular location: Cytoplasm, cytoskeleton : P00491
• Total number of polymer chains: 3
• Total formula weight: 106922.26

Authors

Sawaya, M.R.,Afshar, S. (deposition date: 2009-02-19, release date: 2009-04-14, Last modification date: 2023-09-06)

Primary citation

Afshar, S.,Sawaya, M.R.,Morrison, S.L.
Structure of a mutant human purine nucleoside phosphorylase with the prodrug, 2-fluoro-2'-deoxyadenosine and the cytotoxic drug, 2-fluoroadenine.
Protein Sci., 18:1107-1114, 2009

PubMed Abstract: A double mutant of human purine nucleoside phosphorylase (hDM) with the amino acid mutations Glu201Gln:Asn243Asp cleaves adenosine-based prodrugs to their corresponding cytotoxic drugs. When fused to an anti-tumor targeting component, hDM is targeted to tumor cells, where it effectively catalyzes phosphorolysis of the prodrug, 2-fluoro-2'-deoxyadenosine (F-dAdo) to the cytotoxic drug, 2-fluoroadenine (F-Ade). This cytotoxicity should be restricted only to the tumor microenvironment, because the endogenously expressed wild type enzyme cannot use adenosine-based prodrugs as substrates. To gain insight into the interaction of hDM with F-dAdo, we have determined the crystal structures of hDM with F-dAdo and F-Ade. The structures reveal that despite the two mutations, the overall fold of hDM is nearly identical to the wild type enzyme. Importantly, the residues Gln201 and Asp243 introduced by the mutation form hydrogen bond contacts with F-dAdo that result in its binding and catalysis. Comparison of substrate and product complexes suggest that the side chains of Gln201 and Asp243 as well as the purine base rotate during catalysis possibly facilitating cleavage of the glycosidic bond. The two structures suggest why hDM, unlike the wild-type enzyme, can utilize F-dAdo as substrate. More importantly, they provide a critical foundation for further optimization of cleavage of adenosine-based prodrugs, such as F-dAdo by mutants of human purine nucleoside phosphorylase.

PubMed: 19388075
DOI: 10.1002/pro.91

Experimental method

X-RAY DIFFRACTION (2.3 Å)