Vinylphosphonate internucleotide linkages inhibit the activity of PcrA DNA helicase

During the past 5 years a great deal of structural and biochemical information has given us a detailed insight into the molecular mechanism of action of the PcrA DNA helicase and challenged previous notions about the molecular mechanism of action of helicases in general. Despite this wealth of information the mechanisms of the interaction of helicases with their DNA substrates and their unidirectional translocation along ssDNA are poorly understood. In this study, we synthesized a chemically modified DNA substrate with reduced backbone rotational flexibility and minimal steric hindrance and studied its effect on the activity of the monomeric 3'-5' DNA helicase, PcrA. Our results show that a single modification on the backbone of the translocating strand is sufficient to inhibit the activity of PcrA helicase, suggesting that rotational flexibility of the backbone is important for efficient unwinding.