Structure of Type IIL Restriction-Modification Enzyme MmeI in Complex with DNA Has Implications for Engineering New Specificities.

• Main Authors: Scott J Callahan, Yvette A Luyten, Yogesh K Gupta, Geoffrey G Wilson, Richard J Roberts, Richard D Morgan, Aneel K Aggarwal
• Format: Article
• Language: English
• Published: Public Library of Science (PLoS) 2016-04-01
• Series: PLoS Biology
• Online Access: http://europepmc.org/articles/PMC4833311?pdf=render
• ISSN: 1544-9173; 1545-7885

The creation of restriction enzymes with programmable DNA-binding and -cleavage specificities has long been a goal of modern biology. The recently discovered Type IIL MmeI family of restriction-and-modification (RM) enzymes that possess a shared target recognition domain provides a framework for engineering such new specificities. However, a lack of structural information on Type IIL enzymes has limited the repertoire that can be rationally engineered. We report here a crystal structure of MmeI in complex with its DNA substrate and an S-adenosylmethionine analog (Sinefungin). The structure uncovers for the first time the interactions that underlie MmeI-DNA recognition and methylation (5'-TCCRAC-3'; R = purine) and provides a molecular basis for changing specificity at four of the six base pairs of the recognition sequence (5'-TCCRAC-3'). Surprisingly, the enzyme is resilient to specificity changes at the first position of the recognition sequence (5'-TCCRAC-3'). Collectively, the structure provides a basis for engineering further derivatives of MmeI and delineates which base pairs of the recognition sequence are more amenable to alterations than others.