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PLoS Biol. 2016 Apr 15;14(4):e1002442. doi: 10.1371/journal.pbio.1002442. eCollection 2016 Apr.

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

Author information

1
Department of Structural and Chemical Biology, Mount Sinai School of Medicine, New York, New York, United States of America.
2
New England Biolabs Inc., Ipswich, Massachusetts, United States of America.

Abstract

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.

PMID:
27082731
PMCID:
PMC4833311
DOI:
10.1371/journal.pbio.1002442
[Indexed for MEDLINE]
Free PMC Article

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