Estimated pKa values for specific amino acid residues in daptomycin

Daptomycin is a cyclic lipopeptide antibiotic. The ionization constants of daptomycin have not been individually elucidated. The objective of this research is to determine the sequence‐specific ionization constants of daptomycin in the monomeric state. The pH titrations of daptomycin were performed by nuclear magnetic resonance (NMR) spectroscopy. The sequence‐specific pKa values for the four acidic residues and one aromatic amine (Kyn‐13) in daptomycin were determined by two‐dimensional total correlation spectroscopy 1H NMR. From the NMR pH titration, the estimated pKa values for Asp‐3, Asp‐9, and methylglutamic acid (mGlu‐12) were determined to be 4.2, 3.8, and 4.6 in the absence of salt, and 4.1, 3.8, and 4.4 in the presence of 150 mM NaCl, respectively. The pKa value for Asp‐7 is estimated to be approximately 1.0 in the absence of salt and 1.3 in the presence of salt. The estimated Hill coefficients for Asp‐7 were 0.72 and 1.31 in the absence and presence of salt, respectively. The increase in Hill coefficients from 0.72 to 1.31 with increasing salt concentration is consistent with the estimated lower pKa in the absence of salt, and suggests that a salt bridge is formed in solution possibly between Asp‐7 acidic group and the neighboring Orn‐6 basic group. © 2011 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 100:4225-4233, 2011