%0 Journal Article
%T The crystal structures of Sau3AI with and without bound DNA suggest a self-activation-based DNA cleavage mechanism.
%A Liu Y
%A Xu C
%A Zhou H
%A Wang W
%A Liu B
%A Li Y
%A Hu X
%A Yu F
%A He J
%J Structure
%V 31
%N 11
%D 2023 11 2
%M 37652002
%F 5.871
%R 10.1016/j.str.2023.08.005
%X The type II restriction endonuclease Sau3AI cleaves the sequence 5'-GATC-3' in double-strand DNA producing two sticky ends. Sau3AI cuts both DNA strands regardless of methylation status. Here, we report the crystal structures of the active site mutant Sau3AI-E64A and the C-terminal domain Sau3AI-C with a bound GATC substrate. Interestingly, the catalytic site of the N-terminal domain (Sau3AI-N) is spatially blocked by the C-terminal domain, suggesting a potential self-inhibition of the enzyme. Interruption of Sau3AI-C binding to substrate DNA disrupts Sau3AI function, suggesting a functional linkage between the N- and C-terminal domains. We propose that Sau3AI-C behaves as an allosteric effector binding one GATC substrate, which triggers a conformational change to open the N-terminal catalytic site, resulting in the subsequent GATC recognition by Sau3AI-N and cleavage of the second GATC site. Our data indicate that Sau3AI and UbaLAI might represent a new subclass of type IIE restriction enzymes.