Spectroscopic glimpses of the transition state of ATP hydrolysis trapped in a bacterial DnaB helicase

Here, the authors use solid-state NMR and EPR measurements to characterise the ATP hydrolysis transition state of the oligomeric bacterial DnaB helicase from Helicobacter pylori, which was trapped by using aluminium fluoride as a chemical mimic. They identify protein protons that coordinate to the phosphate groups of ADP and DNA and observe that the aluminium fluoride unit is highly mobile and fast-rotating. The ATP hydrolysis transition state of motor proteins is a weakly populated protein state that can be stabilized and investigated by replacing ATP with chemical mimics. We present atomic-level structural and dynamic insights on a state created by ADP aluminum fluoride binding to the bacterial DnaB helicase from Helicobacter pylori. We determined the positioning of the metal ion cofactor within the active site using electron paramagnetic resonance, and identified the protein protons coordinating to the phosphate groups of ADP and DNA using proton-detected P-31,H-1 solid-state nuclear magnetic resonance spectroscopy at fast magic-angle spinning > 100 kHz, as well as temperature-dependent proton chemical-shift values to prove their engagements in hydrogen bonds. F-19 and Al-27 MAS NMR spectra reveal a highly mobile, fast-rotating aluminum fluoride unit pointing to the capture of a late ATP hydrolysis transition state in which the phosphoryl unit is already detached from the arginine and lysine fingers.

Automated summary

The authors utilized solid-state NMR and EPR measurements to investigate the ATP hydrolysis transition state of the oligomeric bacterial DnaB helicase from Helicobacter pylori, with aluminium fluoride as a chemical mimic. They identified protein protons coordinating to the phosphate groups of ADP and DNA, highlighting the highly mobile and fast-rotating nature of the aluminium fluoride unit. This study provides insights into a state formed by ADP aluminum fluoride binding to the bacterial DnaB helicase, with detailed information on metal ion cofactor positioning and protein protons involvement in hydrogen bonds.