Multi-subunit RNA polymerases, which synthesize an RNA polymer from a DNA template, are highly conserved in evolution and are highly dynamic. Because RNA polymerases move along a DNA template as RNA is synthesized, they are considered to be 'molecular motors', moving in single base increments. RNA polymerases utilize four nucleoside triphosphate substrates to incorporate bases into the growing RNA polymer. During each base addition, the substrate added is specified by the sequence of the DNA template. Supercomputer simulation is used to understand the atomic motions of RNA polymerase during RNA polymerization and during RNA polymerase translocation. In order to better simulate the polymerization reaction and RNA polymerase movement along template DNA, different stages of the reaction are simulated separately. To merge consecutive steps in the reaction, a method termed 'replica exchange' is used in which the likeliest trajectory between two reaction steps is simulated. Because simulations are done in a bath of water and ions, predictions are obtained for the likeliest water placements in the structure. During the polymerization reaction, RNA polymerases exclude water. A major effort is to learn the importance of water exclusion in polymerization and in maintaining the accuracy of polymerization. Many amino acid changes have been made in RNA polymerases resulting in altered protein function. To better understand these changes, altered proteins will be studied for their polymerization and translocation functions, and then analyzed by computer simulation to ensure that results from chemical tests are supported by predicted atomic motions.
Broader impacts Multi-subunit RNA polymerases are necessary for accurate genetic information flow in biological systems. As such, the mechanism, dynamics and accuracy of RNA synthesis are fundamental to complex living systems. Sophisticated computer simulation is applied to one of the largest and most difficult problems in modern biology. Trainees will become expert in biochemical, genetic and computational methods, preparing them as flexible, interdisciplinary scientists. Computer simulation data will be incorporated into undergraduate teaching at Michigan State University. Every effort will be made to recruit talented undergraduates and underrepresented groups into this research program, for instance, through the MSU IDEAS research program (www.bmb.msu.edu/~ugrad/research/res-oppor-minority.htm ).
