TY - JOUR
T1 - Mutations of Asp540 and the domain-connecting residues synergistically enhance Pyrococcus furiosus DNA ligase activity
AU - Tanabe, Maiko
AU - Ishino, Sonoko
AU - Ishino, Yoshizumi
AU - Nishida, Hirokazu
N1 - Funding Information:
The authors thank Prof. K. Morikawa (Kyoto Univ.) and Drs. H. Kambara and M. Shirai (Hitachi, Ltd.) for their support. Y.I. was supported by Grants-in-Aid from the Ministry of Education, Culture, Sports, Science and Technology of Japan (Grant numbers 21113005 and 23310152 ).
PY - 2014/1/21
Y1 - 2014/1/21
N2 - The structure of Pyrococcus furiosus DNA ligase (PfuLig), which architecturally resembles human DNA ligase I (hLigI), revealed that the C-terminal helix stabilizes the closed conformation through several ionic interactions between two domains (adenylylation domain (AdD) and C-terminal OB-fold domain (OBD)). This helix is oriented differently in DNA-bound hLigI, suggesting that the disruption of its interactions with AdD facilitates DNA binding. Previously, we demonstrated that the replacement of Asp540 with arginine improves the ligation activity. Here we report that the combination of the Asp540-replacement and the elimination of ionic residues in the helix, forming interactions with AdD, effectively enhanced the activity.
AB - The structure of Pyrococcus furiosus DNA ligase (PfuLig), which architecturally resembles human DNA ligase I (hLigI), revealed that the C-terminal helix stabilizes the closed conformation through several ionic interactions between two domains (adenylylation domain (AdD) and C-terminal OB-fold domain (OBD)). This helix is oriented differently in DNA-bound hLigI, suggesting that the disruption of its interactions with AdD facilitates DNA binding. Previously, we demonstrated that the replacement of Asp540 with arginine improves the ligation activity. Here we report that the combination of the Asp540-replacement and the elimination of ionic residues in the helix, forming interactions with AdD, effectively enhanced the activity.
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U2 - 10.1016/j.febslet.2013.10.037
DO - 10.1016/j.febslet.2013.10.037
M3 - Article
C2 - 24211832
AN - SCOPUS:84892365051
VL - 588
SP - 230
EP - 235
JO - FEBS Letters
JF - FEBS Letters
SN - 0014-5793
IS - 2
ER -