Specificity in protein-DNA interactions: energetic recognition by the (cytosine-C5)-methyltransferase from HhaI

Sequence-specific interactions between proteins and DNA are essential for a variety of biological functions. The (cytosine-C5)-methyltransferase from HhaI (M.HhaI) specifically modifies the second base in GCGC sequences, employing a base flipping mechanism to access the target base being chemically modified. The mechanism of sequence-specific recognition of M.HhaI is not evident based on crystallographic structures, leading to the suggestion that recognition is linked to the flipping event itself, a process that may be referred to as energetic recognition. Using computational methods, it is shown that the free energy barriers to flipping are significantly higher in non-cognate versus the cognate sequence, supporting the energetic recognition mechanism. Energetic recognition is imparted by two protein "selectivity filters" that function via a "web" of protein-DNA interactions in short-lived, high energy states present along the base flipping pathway. Other sequence-specific DNA binding proteins whose function involves significant distortion of DNA's conformation may use a similar recognition mechanism.