N-Terminus and Lysine Side Chain pKa Values of Melittin in Aqueous Solutions and Micellar Dispersions Measured by 15N NMR

Lingyang Zhu, Marvin D. Kemple, Peng Yuan, Franklyn G. Prendergast

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41 Scopus citations


Melittin (MLT) is a 26-amino acid cytolytic peptide from the Apis mellifera honey bee. It is known to exist as an α-helical tetramer, as an α-helical monomer, or as a monomeric random coil depending on solvent conditions. The charge state of MLT is believed to be a major factor in determining its aggregation properties and its interaction with lipids. Several, contradictory, indirect measurements of the pKa values of the three lysine groups in MLT have been reported. In the present study, high-resolution 15N NMR at 50.6 MHz was used to directly measure the pKa values of the amino groups of the Gly-1, Lys-7, Lys-21, and Lys-23 residues of MLT. Specifically, the pH dependence of MLT 15N chemical shifts was measured separately for the isotopically enriched backbone nitrogen of Gly-1 and the side chain nitrogen atoms of Lys-7, Lys-21, and Lys-23 at a MLT concentration of 1.2 mM and a temperature of 23 °C. Measurements were made for MLT in potassium phosphate buffer, in neat water, and in l-myristoyl-2-hydroxyl-sn-glycero-3-phosphocholine (MMPC) lipid micelles. The experiments showed for MLT tetramer in aqueous phosphate buffer that the amino nitrogen of Gly-1 has a pKa of 8.15, and that the Lys-7, Lys-21, and Lys-23 side chain nitrogen atoms have pKa values of 10.21, 10.03, and 10.24 respectively. The pKa values were somewhat lower for MLT in neat water with Gly-1 at 7.85 (in which case MLT is a monomer), and Lys-21 and Lys-23 at 9.83 and 9.70, respectively (in which case MLT is a tetramer). Similarly for MLT in 48 mM MMPC micelles and 50 mM potassium phosphate, the values are 7.90, 10.09, 9.87, and 10.10 for Gly-1, Lys-7, Lys-21, and Lys-23, respectively. These pKa values indicate that the lysine residues are positively charged when MLT forms a tetramer at pH values between 8.5 and 10.

Original languageEnglish (US)
Pages (from-to)13196-13202
Number of pages7
Issue number40
StatePublished - Oct 1995

ASJC Scopus subject areas

  • Biochemistry


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