We are using our single molecule techniques to study the human transcription factor IIH. TFIIH is a large, multi-domain protein that acts on DNA in two distinct ways. A translocase domain pushes DNA toward the RNA polymerase active site during transcription initiation, creating a bubble in the DNA by inducing torsional strain. A separate helicase domain unwinds DNA around a damage site to enable access by nucleotide excision repair machinery. Much is still unknown about how it interacts with DNA and how it negotiates its roles in RNA transcription, DNA repair, and the coupling of these two pathways. Single molecule methods offer the opportunity to understand these mechanisms further by following the dynamic molecular processes in real-time.

Type IA topoisomerases relieve torsional strain in DNA by cleaving the backbone of one strand of a DNA duplex, passing the other strand through the break, and re-sealing the DNA. This process relaxes supercoiled DNA in a controlled step-wise manner. Because this process is critical to genome integrity, these enzymes are ideal targets for novel antibiotics. Our lab is using a combination of magnetic tweezers, fluorescence, and MD simulations to characterize the type IA topoisomerases of M. smegmatis and M. tuberculosis.