QM/MM Study of the Autophosphorylation Reaction in Histidine Kinases

ADRIAN TURJANSKI; OSVALDO BURASTERO; MARCELO MARTI; LUCAS A. DEFELIPE

Buenos Aires

Congreso; ISCB Latin America; 2016

Asociación argentina de bioinformática y biología computacional

Two component systems (TCSs) comprise signal-transduction pathways which respond to stimulus detected by Histidine-Kinase (HK) type sensors. Recognition causes autophosporylation of the HK and subsequent transfer of the phosphate group to its cognate receptor that acts as transcriptional regulator. They are potential pharmacological targets because they are ubiquitous among prokaryotes but are totally absent in superior eukaryotes. HKs are homodimeric receptors with a modular structure which simplest form consist of a sensor domain (preferentially extracytoplasmic) and a cytoplasmatic transmisor core. While the former is more variable, the latter is widely conserved. The autophosphorylation reaction implies the transference from the ATP γ-phosphate to a conserved histidine to form a highly unstable N-P bond. It?s mechanism remains elusive due to the great flexibility of the active site and the lack of structural information until recent years. For this reason, in this work we proposed to resolve it using the CpxA HK as model and using QM/MM hybrid technics. The first step was to evaluate different quantum level approximations to describe the phosphorylated histidine and evaluate the product stability in different protonation states. We establish that DFT describes the system correctly and which is the more stable product. In a second step we analyze a set of possible reaction mechanisms utilizing free energy profiles made by MSMD and Jarzynski equality. To conclude, we find a concerted one-step mechanism with: the histidine protonated in δ stabilized by a glutamic; a plane transition state and a 15 Kcal/mol activation barrier.