Evidence for a functional dimeric form of the PcrA helicase in DNA unwinding.
Evidence for a functional dimeric form of the PcrA helicase in DNA unwinding.
复制标题
DNA 解旋中 PcrA 解旋酶功能性二聚体形式的证据
DOI：
10.1093/nar/gkm1174
复制
发表时间：
2008-04
影响因子：
14.9
通讯作者：
Xi XG
中科院分区：
文献类型：
--
作者：
Yang Y;Dou SX;Ren H;Wang PY;Zhang XD;Qian M;Pan BY;Xi XG
PcrA helicase, a member of the superfamily 1, is an essential enzyme in many bacteria. The first crystal structures of helicases were obtained with PcrA. Based on structural and biochemical studies, it was proposed and then generally believed that PcrA is a monomeric helicase that unwinds DNA by an inchworm mechanism. But a functional state of PcrA from unwinding kinetics studies has been lacking. In this work, we studied the kinetic mechanism of PcrA-catalysed DNA unwinding with fluorometric stopped-flow method under both single- and multiple-turnover conditions. It was found that the PcrA-catalysed DNA unwinding depended strongly on the PcrA concentration as well as on the 3′-ssDNA tail length of the substrate, indicating that an oligomerization was indispensable for efficient unwinding. Study of the effect of ATP concentration on the unwinding rate gave a Hill coefficient of ∼2, suggesting strongly that PcrA functions as a dimer. It was further determined that PcrA unwound DNA with a step size of 4 bp and a rate of ∼9 steps per second. Surprisingly, it was observed that PcrA unwound 12-bp duplex substrates much less efficiently than 16-bp ones, highlighting the importance of protein-DNA duplex interaction in the helicase activity. From the present studies, it is concluded that PcrA is a dimeric helicase with a low processivity in vitro. Implications of the experimental results for the DNA unwinding mechanism of PcrA are discussed.
登录
查看更多内容
影响因子：
4.8
作者：
Dou, SX;Wang, PY;Xi, XG
通讯作者：
Xi, XG
影响因子：
5.7
作者：
Kim, JL;Morgenstern, KA;Caron, PR
通讯作者：
Caron, PR
影响因子：
56.9
作者：
Ali, JA;Lohman, TM
通讯作者：
Lohman, TM
影响因子：
5.6
作者：
Cheng, W;Hsieh, J;Lohman, TM
通讯作者：
Lohman, TM
DOI：
10.1073/pnas.242479399
发表时间：
2002-12-10
影响因子：
11.1
作者：
Cheng, W;Brendza, KM;Lohman, TM
通讯作者：
Lohman, TM