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Optical Control of DNA Helicase Function through Genetic Code Expansion
Author(s) -
Luo Ji,
Kong Muwen,
Liu Lili,
Samanta Subhas,
Van Houten Bennett,
Deiters Alexander
Publication year - 2017
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201600624
Subject(s) - helicase , dna repair , dna , biology , nucleotide excision repair , atpase , microbiology and biotechnology , biochemistry , circular bacterial chromosome , biophysics , genetics , enzyme , dna replication , gene , rna
Nucleotide excision repair (NER) is a general DNA repair mechanism that is capable of removing a wide variety of DNA lesions induced by physical or chemical insults. UvrD, a member of the helicase SF1 superfamily, plays an essential role in bacterial NER by unwinding the duplex DNA in the 3′ to 5′ direction to displace the lesion‐containing strand. In order to achieve conditional control over NER, we generated a light‐activated DNA helicase. This was achieved through a site‐specific incorporation of a genetically encoded hydroxycoumarin lysine at a crucial position in the ATP‐binding pocket of UvrD. The resulting caged enzyme was completely inactive in several functional assays. Moreover, enzymatic activity of the optically triggered UvrD was comparable to that of the wild‐type protein, thus demonstrating excellent OFF to ON switching of the helicase. The developed approach provides optical control of NER, thereby laying a foundation for the regulation of ATP‐dependent helicase functions in higher organisms. In addition, this methodology is applicable to the light‐activation of a wide range of ATPases.