Kooistra J, Haijema B J, Hesseling-Meinders A, Venema G
Department of Genetics, University of Groningen, Haren, The Netherlands.
Mol Microbiol. 1997 Jan;23(1):137-49. doi: 10.1046/j.1365-2958.1997.1991570.x.
Various mutations were introduced in a conserved helicase domain (motif VI) of the AddA subunit of the Bacillus subtilis ATP-dependent nuclease (AddAB) by site-directed mutagenesis. These mutations affected the helicase activity and the ATP-dependent exonuclease activity on double-stranded DNA (dsDNA) as the substrate to various degrees, but had hardly any effect on the exonuclease activity on single-stranded DNA (ssDNA), suggesting that exonuclease activity on dsDNA of the enzyme requires unwinding of the DNA. This idea was supported by the finding that, initially, the rate and extent of unwinding of the DNA were higher than those of its degradation to acid-soluble products by the exonucleolytic activity. The effects of the mutations on DNA repair and recombination correlated strongly with their effects on helicase activity. Taken together, these results suggest that motif VI is essential for the helicase activity, and that this activity is required for DNA repair and recombination.
通过定点诱变，在枯草芽孢杆菌ATP依赖性核酸酶（AddAB）的AddA亚基的保守解旋酶结构域（基序VI）中引入了各种突变。这些突变不同程度地影响了解旋酶活性以及以双链DNA（dsDNA）为底物的ATP依赖性核酸外切酶活性，但对单链DNA（ssDNA）上的核酸外切酶活性几乎没有影响，这表明该酶对dsDNA的核酸外切酶活性需要DNA解旋。这一观点得到以下发现的支持：最初，DNA解旋的速率和程度高于其通过核酸外切活性降解为酸溶性产物的速率和程度。这些突变对DNA修复和重组的影响与其对解旋酶活性的影响密切相关。综上所述，这些结果表明基序VI对解旋酶活性至关重要，并且这种活性是DNA修复和重组所必需的。
