Halgasova Nora, Krajcikova Daniela, Kraus Daniel, Bukovska Gabriela
Department of Genomics and Biotechnology, Institute of Molecular Biology, Slovak Academy of Sciences, Dubravska cesta 21, 845 51, Bratislava, Slovakia.
Department of Microbial Genetics, Institute of Molecular Biology, Slovak Academy of Sciences, Dubravska cesta 21, 845 51, Bratislava, Slovakia.
Virology. 2021 Jun;558:96-109. doi: 10.1016/j.virol.2021.02.016. Epub 2021 Mar 11.
The multifunctional phage replication protein gp43 is composed of an N-terminal prim-pol domain and a C-terminal domain similar to the SF4-type replicative helicases. We prepared four mutants all missing the prim-pol domain with the helicase core flanked by accessory N- and C-terminal regions truncated to varying extents. The shortest fragment still possessing strong ssDNA-dependent ATPase activity and helicase activity was gp43HEL519-983. The other proteins tested were gp43HEL557-983, gp43HEL519-855 and gp43HEL519-896. Removal of the 38 N-terminal residues in gp43HEL557-983, or the 128 and 87 C-terminal residues in gp43HEL519-855 and gp43HEL519-896, resulted in a significant decrease in the ATPase activities. The 38-amino acid N-terminal region has probably a function in modulating DNA binding and protein oligomerization. Deletion of the 87 C-terminal residues resulted in a twofold increase in the unwinding rate. This region is likely indispensable for binding to DNA substrates.
多功能噬菌体复制蛋白gp43由一个N端引物-聚合酶结构域和一个与SF4型复制解旋酶相似的C端结构域组成。我们制备了四个均缺失引物-聚合酶结构域的突变体，其解旋酶核心两侧的N端和C端辅助区域被不同程度截短。仍具有较强单链DNA依赖性ATP酶活性和解旋酶活性的最短片段是gp43HEL519-983。测试的其他蛋白是gp43HEL557-983、gp43HEL519-855和gp43HEL519-896。gp43HEL557-983中38个N端残基的缺失，或gp43HEL519-855和gp43HEL519-896中128个和87个C端残基的缺失，导致ATP酶活性显著降低。38个氨基酸的N端区域可能具有调节DNA结合和蛋白质寡聚化的功能。87个C端残基的缺失导致解旋速率增加两倍。该区域可能对于与DNA底物的结合不可或缺。
