The preprocessed initial files that feed the molecular dynamics (MD) simulation packages dramatically influence the outcome of the simulations. However, the popular MD simulation packages depend, to a great extent, on the user’s experience in the preparation of MD simulation systems. In this work, we present an easy-to-use tool called MDBuilder, a PyMOL plugin that assists researchers in building the starting structures for multiple popular MD simulation packages. MDBuilder is not only designed to assist MD beginners to overcome the steep learning curve by providing a menu-oriented, point-and-click user graphic interface (GUI), but also to provide an alternative way to prepare the input files for some highly scalable CHARMM force field-based MD simulation packages. The platform-independent GUI is implemented as a PyMOL plugin using the Python language, and it has been tested on Windows and Linux platforms. The source code and documentation of MDBuilder can be downloaded freely from https://github.com/HuiLiuCode/MDBuilder under the GNU General Public License.

CRISPR-Cas9 systems constitute bacterial adaptive immune systems that protect against phage infections. Bacteriophages encode anti-CRISPR proteins (Acrs) that mitigate the bacterial immune response. However, the structural basis for their inhibitory actions from a molecular perspective remains elusive. In this study, through microsecond atomistic molecular dynamics simulations, we demonstrated the remarkable flexibility of Streptococcus pyogenes Cas9 (SpyCas9) and its conformational adaptability during interactions with AcrIIA4 and AcrIIA2. Specifically, we demonstrated that the binding of AcrIIA4 and AcrIIA2 to SpyCas9 induces a conformational rearrangement that causes spatial separation between the nuclease and cleavage sites, thus making the endonuclease inactive. This separation disrupts the transmission of signals between the protospacer adjacent motif recognition and nuclease domains, thereby impeding the efficient processing of double-stranded DNA. The simulation also reveals that AcrIIA4 and AcrIIA2 cause different structural variations of SpyCas9. Our research illuminates the precise mechanisms underlying the suppression of SpyCas9 by AcrIIA4 and AcrIIA2, thus presenting new possibilities for controlling genome editing with higher accuracy.