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Lysine Acetylation of Nuclear Pif1 Regulates Its Helicase Function
Author(s) -
Oye Onyekachi E,
Sausen Christopher W,
Bochman Matthew W,
Balakrishnan Lata
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.lb51
Subject(s) - acetylation , helicase , acetyltransferase , lysine , rna helicase a , biochemistry , biology , chemistry , microbiology and biotechnology , rna , gene , amino acid
Petite integration frequency 1 (Pif1) is a 5′‐3′ helicase which plays a significant role in maintaining the fidelity of nuclear and mitochondrial DNA. Pif1 preferentially unwinds RNA‐DNA hybrids during key metabolic processes such as DNA replication, repair and telomere maintenance. Regulation of Pif1 activity by phosphorylation modulates the role of this helicase in telomere maintenance. Our current work is focused on understanding how lysine acetylation modifies the protein's activity. Lysine acetylation of proteins is a dynamic modification that is regulated by the activity of acetyltransferases and deacetylases. Using biochemical assays we characterized the enzymatic activity of acetylated Pif1, which was modified in vitro using the Gcn5/Ada2/Ada3 and Rtt109/Vps75 acetyltransferase complexes. Helicase assays revealed an enhanced unwinding function of Pif1 upon protein acetylation. Additionally, the ATPase activity of acetylated Pif1 was also significantly stimulated compared to the unmodified form. In concert, the binding function of acetylated Pif1 to its cognate substrates was also improved when assessed using gel electro‐mobility shift assay (EMSA) in tandem with Bio‐layer interferometry technology (BLI). Mass spectrometric analysis of the in vitro acetylated Pif1 protein revealed some acetylated lysine residues on the N‐terminus of the protein. Current work is focused on understanding how acetylation based Pif1 regulation impacts its role in the maintenance of genome fidelity. Support or Funding Information IUPUI New Faculty Start Up Grant NIH R00 GM98328