4JST

Structure of Clostridium thermocellum polynucleotide kinase bound to UTP

Summary for 4JST

• Entry DOI: 10.2210/pdb4jst/pdb
• Related: 4GP6 4GP7 4JSY 4JT2 4JT4
• Descriptor: Metallophosphoesterase, MAGNESIUM ION, URIDINE 5'-TRIPHOSPHATE, ... (5 entities in total)
• Functional Keywords: rna repair, p-loop phosphotransferase, polynucleotide kinase, transferase
• Biological source: Clostridium thermocellum
• Total number of polymer chains: 2
• Total formula weight: 40362.01

Authors

Das, U.,Wang, L.K.,Smith, P.,Shuman, S. (deposition date: 2013-03-22, release date: 2013-08-28, Last modification date: 2024-11-06)

Primary citation

Das, U.,Wang, L.K.,Smith, P.,Shuman, S.
Structural and biochemical analysis of the phosphate donor specificity of the polynucleotide kinase component of the bacterial pnkphen1 RNA repair system.
Biochemistry, 52:4734-4743, 2013

PubMed Abstract: Clostridium thermocellum Pnkp is the end-healing and end-sealing subunit of a bacterial RNA repair system. CthPnkp is composed of three catalytic modules: an N-terminal 5'-OH polynucleotide kinase, a central 2',3' phosphatase, and a C-terminal ligase. The crystal structure of the kinase domain bound to ATP•Mg(2+) revealed a rich network of ionic and hydrogen-bonding contacts to the α, β, and γ phosphates. By contrast, there are no enzymic contacts to the ribose and none with the adenine base other than a π-cation interaction with Arg116. Here we report that the enzyme uses ATP, GTP, CTP, UTP, or dATP as a phosphate donor for the 5'-OH kinase reaction. The enzyme also catalyzes the reverse reaction, in which a polynucleotide 5'-PO4 group is transferred to ADP, GDP, CDP, UDP, or dADP to form the corresponding NTP. We report new crystal structures of the kinase in complexes with GTP, CTP, UTP, and dATP in which the respective nucleobases are stacked on Arg116 but make no other enzymic contacts. Mutating Arg116 to alanine elicits a 10-fold increase in Km for ATP but has little effect on kcat. These findings illuminate the basis for nonspecific donor nucleotide utilization by a P-loop phosphotransferase.

PubMed: 23721485
DOI: 10.1021/bi400412x

Experimental method

X-RAY DIFFRACTION (2.03 Å)