TIME EVOLUTION OF THE ALLOSTERIC TRANSITION OF ASPARTATE TRANSCARBAMOYLASE

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Aspartate transcarbamoylase from E. coli exits in two conformational states, a low-activity low-affinity T state and a high-activity high-affinity R state. The enzyme not only catalyzes the first reaction in the pyrimidine biosynthesis pathway, but is also involved in the control of the rate of this entire metabolic pathway. Control is thought to be achieved by altering the ratio of the T and R forms. The T and R states of the enzyme are both functionally and structurally distinct. During the conversion of the enzyme from the T to the R state, the enzyme undergoes an elongation of appoximately 11 ¿¿ along with simultaneous rotations of subunits, which can easily be detected by SAXS. We have previously proposed a mechanism for a concerted allosteric transition from the T to the R states. We have been able to perform one set of experiments at SSRL using time-resolved SAXS to directly follow the time course of the structural transition from the T to the R state. These preliminary data suggest that a structural intermediate is formed during the transition. This proposal is for additional beam time at SSRL to continue the investigation into the allosteric mechanism of the enzyme and how the heterotropic effects influence the structural transition from the T to the R state. This will be the first time for an allosteric enzyme that the time evolution of the allosteric structural change will be followed in real time by SAXS.