6OIX

Structure of Escherichia coli dGTPase bound to GTP

6OIX の概要

• エントリーDOI: 10.2210/pdb6oix/pdb
• 分子名称: Deoxyguanosinetriphosphate triphosphohydrolase, GUANOSINE-5'-TRIPHOSPHATE, MANGANESE (II) ION, ... (4 entities in total)
• 機能のキーワード: dntp triphosphohydrolase, metalloenzyme, metal binding protein, xfel, e. coli dgtpase, hydrolase
• 由来する生物種: Escherichia coli (strain K12)
• タンパク質・核酸の鎖数: 6
• 化学式量合計: 360293.89

構造登録者

Barnes, C.O.,Wu, Y.,Calero, G. (登録日: 2019-04-09, 公開日: 2019-05-08, 最終更新日: 2024-03-13)

主引用文献

Barnes, C.O.,Wu, Y.,Song, J.,Lin, G.,Baxter, E.L.,Brewster, A.S.,Nagarajan, V.,Holmes, A.,Soltis, S.M.,Sauter, N.K.,Ahn, J.,Cohen, A.E.,Calero, G.
The crystal structure of dGTPase reveals the molecular basis of dGTP selectivity.
Proc.Natl.Acad.Sci.USA, 116:9333-9339, 2019

PubMed Abstract: Deoxynucleotide triphosphohydrolases (dNTPases) play a critical role in cellular survival and DNA replication through the proper maintenance of cellular dNTP pools. While the vast majority of these enzymes display broad activity toward canonical dNTPs, such as the dNTPase SAMHD1 that blocks reverse transcription of retroviruses in macrophages by maintaining dNTP pools at low levels, ()dGTPase is the only known enzyme that specifically hydrolyzes dGTP. However, the mechanism behind dGTP selectivity is unclear. Here we present the free-, ligand (dGTP)- and inhibitor (GTP)-bound structures of hexameric dGTPase, including an X-ray free-electron laser structure of the free -dGTPase enzyme to 3.2 Å. To obtain this structure, we developed a method that applied UV-fluorescence microscopy, video analysis, and highly automated goniometer-based instrumentation to map and rapidly position individual crystals randomly located on fixed target holders, resulting in the highest indexing rates observed for a serial femtosecond crystallography experiment. Our structures show a highly dynamic active site where conformational changes are coupled to substrate (dGTP), but not inhibitor binding, since GTP locks dGTPase in its apo- form. Moreover, despite no sequence homology, -dGTPase and SAMHD1 share similar active-site and HD motif architectures; however, -dGTPase residues at the end of the substrate-binding pocket mimic Watson-Crick interactions providing guanine base specificity, while a 7-Å cleft separates SAMHD1 residues from dNTP bases, abolishing nucleotide-type discrimination. Furthermore, the structures shed light on the mechanism by which long distance binding (25 Å) of single-stranded DNA in an allosteric site primes the active site by conformationally "opening" a tyrosine gate allowing enhanced substrate binding.

PubMed: 31019074
DOI: 10.1073/pnas.1814999116

実験手法

X-RAY DIFFRACTION (3.15 Å)