Tomoaia-Cotisel, M; Stewart, LC; Kates, M; Zsako, J; Chifu, E; Mocanu, A; Frangopol, PT; Noe, LJ; Quinn, PJ
Acid dissociation constants of 2,3-diphytanyl-sn-glycero-1-phosphoryl-sn-3'-glycero-1'-methylphosphate (PGP-Me), the major phospholipid in extreme halophiles (Halobacteriaceae), and of the demethylated 2,3-diphytanyl-sn-glycero-1-phosphoryl-sn-3'-glycero-1'-phosphate (PGP) and its deoxy analog 2,3-diphytanyl-sn-glycero-1-phosphoryl-1'-(1',3'-propanediol-3'-phosphate) (dPGP), were calculated by an original mathematical procedure from potentiometric titration data in methanol/water (1:1, v/v) and found to be as follows: for PGP-Me (and presumably PGP), pK1=3.00 and pK2=3.61; for PGP, pK3=11.12; and for dPGP, pK1=2.72, pK2=4.09, and pK3=8.43. High-resolution 31P NMR spectra of intact PGP-Me in benzene/methanol or in aqueous dispersion showed two resonances corresponding to the two P-OH groups, each of which exhibited a chemical shift change in the pH range 2.0-4.5, corresponding to acid dissociation constants pK1=pK2=3.2; no further ionization (pK3) was detected at higher pH values in the range 5-12. The present results show that PGP-Me titrates as a dibasic acid in the pH range 2-8, but above pH 8, it titrates as a tribasic acid, presumably PGP, formed by hydrolysis of the methyl group during the titration with KOH. Calculation of the concentrations of the ionic molecular species of PGP-Me, PGP and dPGP as a function of pH showed that the dianionic species predominate in the pH range 5-9, covering the optimal pH for growth of Halobacteriaceae. The results are consistent with the concept that the hydroxyl group of the central glycerol moiety in PGP-Me and PGP is involved in the formation of an intramolecular hydrogen-bonded cyclic structure of the polar headgroup, which imparts greater stability to the dianionic form of PGP-Me and PGP in the pH range 5-9 and facilitates lateral proton conduction by a process of diffusion along the membrane surface of halobacterial cells. Copyright (C) 1999 Elsevier Science Ireland Ltd.