title: Improved Sensitivity for Long-Distance Measurements in Biomolecules: Five-Pulse Double Electron-Electron Resonance.
abstract: We describe significantly improved long-distance measurements in biomolecules by use of the new multipulse double electron-electron spin resonance (DEER) illustrated with the example of a five-pulse DEER sequence. In this sequence, an extra pulse at the pump frequency is used compared with standard four-pulse DEER. The position of the extra pulse is fixed relative to the three pulses of the detection sequence. This significantly reduces the effect of nuclear spin-diffusion on the electron-spin phase relaxation, thereby enabling longer dipolar evolution times that are required to measure longer distances. Using spin-labeled T4 lysozyme at a concentration less than 50 μM, as an example, we show that the evolution time increases by a factor of 1.8 in protonated solution and 1.4 in deuterated solution to 8 and 12 μs, respectively, with the potential to increase them further. This enables a significant increase in the measurable distances, improved distance resolution, or both.
authors.affiliation: Department of Chemistry and Chemical Biology,  Cornell University , Ithaca, New York 14853, United States.
authors.forename: Peter P
authors.initials: PP
authors.lastname: Borbat
authors.forename: Elka R
authors.initials: ER
authors.lastname: Georgieva
authors.forename: Jack H
authors.initials: JH
authors.lastname: Freed