Unravelling the mechanism of RNA-polymerase forward motion by using mechanical force.
Polymerases form a class of enzymes that act as molecular motors as they move along their nucleic acid substrate during catalysis, incorporating nucleotide triphosphates at the end of the growing chain and consuming chemical energy. A debated issue is how the enzyme converts chemical energy into motion [J. Gelles and R. Landick, Cell 93, 13 (1998)]. In a single molecule assay, we studied how an opposing mechanical force affects the translocation rate of T7 RNA polymerase. Our measurements show that force acts as a competitive inhibitor of nucleotide binding. This result is interpreted in the context of possible models, and with respect to published crystal structures of T7 RNA polymerase. The transcribing complex appears to utilize only a small fraction of the energy of hydrolysis to perform mechanical work, with the remainder being converted to heat.
Thomen
Philippe
P
Laboratoire Pierre Aigrain, Ecole Normale Supérieure, 24 rue Lhomond, 75005 Paris, France.
Lopez
Pascal J
PJ
Heslot
François
F
D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D001710:Biotin / Q000737:chemistry; D003570:Cytidine Triphosphate / Q000737:chemistry / Q000378:metabolism; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D011487:Protein Conformation; D012822:Silicon Dioxide / Q000737:chemistry; D019809:Streptavidin / Q000737:chemistry; D013816:Thermodynamics; D014544:Uridine Triphosphate / Q000737:chemistry / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000378:metabolism*
D004279:DNA, Viral; D014764:Viral Proteins; D003570:Cytidine Triphosphate; D001710:Biotin; D012822:Silicon Dioxide; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D019809:Streptavidin; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D014544:Uridine Triphosphate