Investigation of finite‐pulse radiofrequency‐driven recoupling methods for measurement of intercarbonyl distances in polycrystalline and membrane‐associated HIV fusion peptide samples

Two finite‐pulse radiofrequency‐driven recoupling (RFDR) methods were compared and applied to the measurement of 3–6 Å 13 CO 13 CO distances in polycrystalline and membrane‐associated HIV fusion peptide (HFP) samples. The RFDR methods were based on π pulses and were relatively straightforward to implement and insensitive to pulse imperfections. The two tested methods were: (i) constant‐time double‐quantum buildup with finite pulses (fpCTDQBU) for which the pulse sequence maintained a constant transverse relaxation period while allowing a variable period of dipolar dephasing; and (ii) constant‐time finite‐pulse rf‐driven recoupling (fpRFDR‐CT) for which the duration of transverse relaxation increased with increasing dephasing period. The fpRFDR‐CT method yielded higher signal‐to‐noise and an accurate determination of a ∼5 Å intercarbonyl distance was made in a crystalline peptide which had T 2 ≈ 55 ms. In some contrast, the HFP samples had T 2 ≈ 15 ms and the fpRFDR‐CT data were dominated by transverse relaxation. Examination of the fpCTDQBU sequence showed: (i) the most rapid signal buildup was obtained with application of one 13 C π pulse per rotor period rather than one 13 C π pulse per multiple rotor periods and (ii) the data were insensitive to ∼15 ppm transmitter offset and to ∼5° variation of π pulse nutation angle. For HFP samples which were 13 CO labeled at a single residue, analyses of the fpCTDQBU data were interpreted with a model of mixed parallel and antiparallel β‐strand arrangements in the N ‐terminal region of HFP and loss of parallel β‐sheet structure in the C ‐terminal region of HFP. Copyright © 2007 John Wiley & Sons, Ltd.