Reversible Sheet–Turn Conformational Change of a Cell-Penetrating Peptide in Lipid Bilayers Studied by Solid-State NMR

Abstract

The membrane-bound conformation of a cell-penetrating peptide, penetratin, is investigated using solid-state NMR spectroscopy. The (13)C chemical shifts of (13)C, (15)N-labeled residues in the peptide indicate a reversible conformational change from beta-sheet at low temperature to coil-like at high temperature. This conformational change occurs for all residues examined between positions 3 and 13, at peptide/lipid molar ratios of 1:15 and 1:30, in membranes with 25-50% anionic lipids, and in both saturated DMPC/DMPG (1,2-dimyristoyl-sn-glycero-3-phosphatidylchloline/1,2-dimyristoyl-sn-glycero-3-phosphatidylglycerol) membranes and unsaturated POPC/POPG (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylglycerol) membranes. Thus, it is an intrinsic property of penetratin. The coil state of the peptide has C-H order parameters of 0.23-0.52 for C(alpha) and C(beta) sites, indicating that the peptide backbone is unstructured. Moreover, chemical shift anisotropy lineshapes are uniaxially averaged, suggesting that the peptide backbone undergoes uniaxial rotation around the bilayer normal. These observations suggest that the dynamic state of penetratin at high temperature is a structured turn instead of an isotropic random coil. The thermodynamic parameters of this sheet-turn transition are extracted and compared to other membrane peptides reported to exhibit conformational changes. We suggest that the function of this turn conformation may be to reduce hydrophobic interactions with the lipid chains and facilitate penetratin translocation across the bilayer without causing permanent membrane damage.

Structure and dynamics of cationic membrane peptides and proteins: Insights from solid‐state NMR

“…[104][105][106] Chemical shifts indicate that in anionic bilayers, penetratin conformation depends on the temperature; in the gel phase the peptide exhibits b-sheet 13 C and 15 N shifts while at physiological temperature it exhibits coil or turn chemical shifts. 107 This unusual sheet!coil conformational change occurs in saturated DMPC/DMPG and unsaturated POPC/ POPG membranes, is reversible with temperature, and occurs at a range of peptide/lipid molar ratios. Thus, it is an intrinsic property of the peptide.…”

“…Thus the hexa-Arg TAT could gain up to À15 kcal/mol favorable free energy. 107 We hypothesize that these two contributions overcome the Born repulsion and account for the facile insertion of this highly charged peptide into the lipid membrane.…”

“…Analysis of the peak intensities of the two states indicated that the sheet!coil transition is entropically driven; a positive enthalpy change of þ0.89 kcal/mol is countered by a positive entropy change of þ3.1 cal/(mol K). 107 At high temperature, the backbone exhibits low order parameters of 0.23-0.52. 107 However, among five examined residues, one residue, Arg10, retained the b-strand torsion angles at high temperature and exhibited a nearly rigid-limit CaHa order parameter (S CH ¼ 0.92).…”

“…107 At high temperature, the backbone exhibits low order parameters of 0.23-0.52. 107 However, among five examined residues, one residue, Arg10, retained the b-strand torsion angles at high temperature and exhibited a nearly rigid-limit CaHa order parameter (S CH ¼ 0.92). 108 This difference suggests that Arg molecules, due to their interactions with lipid phosphates, may stabilize the conformation of the otherwise coil or turn-rich penetratin.…”

Molecular Dynamics of Neutral Polymer Bonding Agent (NPBA) as Revealed by Solid-State NMR Spectroscopy

“…Specifically, an array of techniques, including DIPSHIFT [35,36], 2D WISE [37], SUPER [38], 2 H-NMR [39], REDOR [40,41] and relaxation parameters measurements [42], have been well established to investigate the rigidity or mobility of a variety of functional (bio-)polymers. In recent years, lots of efforts have been made towards better understanding the structure-property relationship of different kinds of polymeric species at the atomic or molecular level using SSNMR.…”