Multiple Locations of Peptides in the Hydrocarbon Core of Gel-Phase Membranes Revealed by Peptide (13)C to Lipid (2)H REDOR Solid-State NMR.
 

Multiple Locations of Peptides in the Hydrocarbon Core of Gel-Phase Membranes Revealed by Peptide (13)C to Lipid (2)H REDOR Solid-State NMR.

Multiple Locations of Peptides in the Hydrocarbon Core of Gel-Phase Membranes Revealed by Peptide (13)C to Lipid (2)H REDOR Solid-State NMR.

Biochemistry. 2014 Dec 22;

Authors: Weliky DP

Abstract
Membrane locations of peptides and proteins are often critical to their functions. Solid-state rotational-echo double-resonance (REDOR) NMR is applied to probe the locations of two peptides via peptide (13)CO to lipid (2)H distance measurements. The peptides are KALP, an ? helical membrane-spanning peptide and HFP, the ? sheet N-terminal fusion peptide of the HIV gp41 fusion protein that plays an important role in HIV/host cell membrane fusion. Both peptides are shown to have at least two distinct locations within the hydrocarbon core of gel-phase membranes. The multiple locations are attributed to snorkeling of lysine sidechains for KALP and to the distribution of antiparallel ? sheet registries for HFP. The relative population of each location is also quantitated. To our knowledge, this is the first clear experimental support of multiple peptide locations within the membrane hydrocarbon core. The present data are for gel-phase membranes but the approach should work for liquid-ordered membranes containing cholesterol and may be applicable to liquid-disordered membranes with appropriate additional analysis to take into account protein and lipid motion. This paper also describes methodological development of (13)CO-(2)H REDOR using the lyophilized I4 peptide which is ? helical and (13)CO-labeled at A9 and (2)H?-labeled at A8. The I4 spins are well-approximated as an ensemble of isolated (13)CO-(2)H spin pairs each separated by 5.0 Å with 37 Hz dipolar coupling. A pulse sequence with rectangular 100 kHz (2)H pulses results in rapid and extensive buildup of REDOR (?S/S0) with dephasing time (?). The buildup is well-fitted by a simple exponential function with rate = 24 Hz and extent close to 1. These parameter values reflect non-radiative transitions between the (2)H spin states during the dephasing period. Each spin pair spends ~2/3 time in the (13)CO-(2)H (m=+1) states and ~1/3 time in the (13)CO-(2)H (m=0) state and contributes to the (?S/S0) buildup during the former but not the latter time segments.


PMID: 25531389 [PubMed - as supplied by publisher]



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