Combining multi-mutant and modular thermodynamic cycles to measure energetic coupling networks in enzyme catalysis.

ABSTRACT: We measured and cross-validated the energetics of networks in Bacillus stearothermophilus Tryptophanyl-tRNA synthetase (TrpRS) using both multi-mutant and modular thermodynamic cycles. Multi-dimensional combinatorial mutagenesis showed that four side chains from this "molecular switch" move coordinately with the active-site Mg²⁺ ion as the active site preorganizes to stabilize the transition state for amino acid activation. A modular thermodynamic cycle consisting of full-length TrpRS, its Urzyme, and the Urzyme plus each of the two domains deleted in the Urzyme gives similar energetics. These dynamic linkages, although unlikely to stabilize the transition-state directly, consign the active-site preorganization to domain motion, assuring coupled vecto

SUBMITTER: Carter CW 

PROVIDER: S-EPMC5272822 | biostudies-literature | 2017 May

REPOSITORIES: biostudies-literature