Role of Glutamate 64 in the Activation of the Prodrug 5-Fluorocytosine by Yeast Cytosine Deaminase

SUMMARYHuman APOBEC3H and homologous single-stranded DNA cytosine deaminases are unique to mammals. These DNA editing enzymes function in innate immunity by restricting the replication of viruses and transposons. Misregulated APOBEC3H also contributes to cancer mutagenesis. Here we address the role of RNA in APOBEC3H regulation. APOBEC3H co-purifies with RNA as an inactive protein, and RNase A treatment yields enzyme preparations with stronger DNA deaminase activity. RNA binding-defective mutants are DNA hypermutators. Chromatography profiles and crystallographic data demonstrate a mechanism in which double-stranded RNA mediates enzyme dimerization. RNA binding is required for APOBEC3H cytoplasmic localization and for packaging into HIV-1 particles and antiviral activity. Related DNA deaminases including other APOBEC3 family members and the antibody gene diversification enzyme AID also bind RNA and are predicted to have a similar RNA binding motif suggesting mechanistic conservation and relevance to innate and adaptive immunity and to multiple diseases.HIGHLIGHTSRNA inhibits human APOBEC3H DNA cytosine deaminase activityRNA binding mutants are DNA hypermutatorsX-ray structure demonstrates an RNA duplex-mediated APOBEC3H dimerization mechanismRNA binding is required for packaging into HIV-1 particles and antiviral activity

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Abstract 5395: Mutant oncolytic Herpes simplex virus expressing yeast cytosine deaminase in combination with 5-FC: Viral replication and prodrug bioactivation kinetics in vivo

10.1158/1538-7445.am2011-5395 ◽

2011 ◽

Author(s):

Suguru Yamada ◽

Toshihiko Kuroda ◽

Bryan C. Fuchs ◽

Xiaoying He ◽

Jeffrey Supko ◽

...

Keyword(s):

Herpes Simplex Virus ◽

Herpes Simplex ◽

Viral Replication ◽

Cytosine Deaminase ◽

Oncolytic Herpes Simplex Virus ◽

Simplex Virus ◽

Kinetics In Vivo ◽

Yeast Cytosine Deaminase

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Microseed matrix screening to improve crystals of yeast cytosine deaminase. Erratum

Acta Crystallographica Section D Biological Crystallography ◽

10.1107/s0907444904005001 ◽

2004 ◽

Vol 60 (4) ◽

pp. 801-801 ◽

Cited By ~ 1

Author(s):

Gregory C. Ireton ◽

Barry L. Stoddard

Keyword(s):

Cytosine Deaminase ◽

Yeast Cytosine Deaminase

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Crystal Structure of tRNA Adenosine Deaminase (TadA) fromAquifex aeolicus

Journal of Biological Chemistry ◽

10.1074/jbc.m414541200 ◽

2005 ◽

Vol 280 (16) ◽

pp. 16002-16008 ◽

Cited By ~ 34

Author(s):

Mitsuo Kuratani ◽

Ryohei Ishii ◽

Yoshitaka Bessho ◽

Ryuya Fukunaga ◽

Toru Sengoku ◽

...

Keyword(s):

Crystal Structure ◽

Adenosine Deaminase ◽

Cytosine Deaminase ◽

Amino Acid Residues ◽

Aquifex Aeolicus ◽

Stem Loop ◽

Dimeric Structure ◽

Docking Model ◽

Tetrameric Structure ◽

Yeast Cytosine Deaminase

The bacterial tRNA adenosine deaminase (TadA) generates inosine by deaminating the adenosine residue at the wobble position of tRNAArg-2. This modification is essential for the decoding system. In this study, we determined the crystal structure ofAquifex aeolicusTadA at a 1.8-Å resolution. This is the first structure of a deaminase acting on tRNA.A. aeolicusTadA has an α/β/α three-layered fold and forms a homodimer. TheA. aeolicusTadA dimeric structure is completely different from the tetrameric structure of yeast CDD1, which deaminates mRNA and cytidine, but is similar to the dimeric structure of yeast cytosine deaminase. However, in theA. aeolicusTadA structure, the shapes of the C-terminal helix and the regions between the β4 and β5 strands are quite distinct from those of yeast cytosine deaminase and a large cavity is produced. This cavity contains many conserved amino acid residues that are likely to be involved in either catalysis or tRNA binding. We made a docking model of TadA with the tRNA anticodon stem loop.

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