Induction of morphological changes in model lipid membranes and the mechanism of membrane disruption by a large scorpion-derived pore-forming peptide

¹ Suntory Institute for Bioorganic Research
² AFMB, CNRS UMR 6098
³ Institute of Biotechnology-UNAM

^* To whom correspondence should be addressed. E-mail: nomura{at}sunbor.or.jp.

Submitted on July 10, 2005
Revised on September 8, 2005
Accepted on 9 September 2005

	Abstract

The membrane disruption mechanism of pandinin 1 (pin1), an antimicrobial peptide isolated from the venom of the African scorpion, was studied using ³¹P, ¹³C, ¹H solid-state and multidimensional solution-state nuclear magnetic resonance (NMR) spectroscopy. A high-resolution NMR solution structure of pin1 showed that the two distinct -helical regions move around the central hinge region, which contains Pro¹⁹. ³¹P NMR spectra of lipid membrane in the presence of pin1, at various temperatures, showed that pin1 induces various lipid phase behaviors depending on the acyl chain length and charge of phospholipids. Notably, it was found that pin1 induced formation of the cubic phase in shorter lipid membranes above T_m. Further, the ¹³C NMR spectra of pin1 labeled at Leu²⁸ under magic angle spinning (MAS) indicated that the motion of pin1 bound to the lipid bilayer was very slow, with a correlation time of the order of 10^-3 s. ³¹P NMR spectra of dispersions of four saturated phosphatidyl-cholines in the presence of three types of pin1 derivatives, [W4A, W6A, W15A]-pin1, pin1(1-18), and pin1(20-44), at various temperatures demonstrated that all three pin1 derivatives have a reduced ability to trigger the cubic phase. ¹³C chemical shift values for pin1(1-18) labeled at Val³, Ala¹⁰, or Ala¹¹ under static or slow MAS conditions indicate that pin1(1-18) rapidly rotates around the average helical axis, and the helical rods are inclined at about 30° to the lipid long axis. ¹³C chemical shift values for pin1(20-44) labeled at Gly²⁵, Leu²⁸, or Ala³¹ under static conditions indicate that pin1(20-44) may be isotropically tumbled. ¹H MAS chemical shift measurements suggest that pin1 is located at the membrane-water interface approximately parallel to the bilayer surface. Solid-state NMR results correlated well with the observed biological activity of pin1 in red blood cells and bacteria.

Key Words: Membrae disruption, Orientation, Solid-state NMR, antimicrobial peptide