2H NMR investigation of the organization and dynamics of polyisoprenols in membranes.

J S De Ropp,F A Troy

doi:10.1016/s0021-9258(17)36310-x

Abstract

The polyisoprenols (PIs) dolichol and undecaprenol function as chemical carriers of glycosyl residues in the membrane-directed synthesis of glycoconjugates in prokaryotic and eukaryotic cells. The molecular details of how these lipid cofactors function is unknown. Presented here are results of deuterium NMR investigations of site specifically 2H-labeled PIs incorporated into model membranes. To complement previous omega-terminal PI labeling schemes, a simple synthesis of head group 2H-labeled PIs is presented in which a PI alcohol is esterified with deuterated acetyl chloride. The 2H-labeled PIs, when incorporated into multilamellar membranes composed of phosphatidylcholine, gave rise to 2H NMR powder patterns interpretable in terms of quadrupole splittings (delta vQ) and spin-lattice relaxation times (T1s). Pure isomers of head group 2H-labeled geraniol (C10) and solanesol (C45) gave rise to single splittings while farnesol (C15) gave rise to two sets of splittings due to cis-trans isomerization at the polar terminal double bond. Membranes containing C45 solanesol exhibited a large isotropic component, indicative of limited partitioning of this poly trans PI into the membrane. T1 measurements revealed high rates of motion for PIs relative to cholesterol in similar membrane hosts and revealed correlation times close to the fatty acyl methyl termini in phosphatidylcholine. The smaller PIs showed higher rates of motion but the T1s of head and tail labels were similar. These data indicate that both ends of the esterified PI molecules see similar environments, probably in the bilayer interior, and suggest that the esterified PIs studied here do not appear to adopt a conventional head group-at-interface orientation of lipids within the bilayer.

Highlights

The polyisoprenols (PIS)dolichol and undecaprenol postulate was that thePISparticipated inactive translocation function as chemical carriers of glycosyl residues in of sugar units across cell membranes
Despite the ubiquitous role of PIS in prokaryotic and eukaryotic glycoconjugate synthesis, and the wealth of information on their distribution, metabolism, and biosynthesis [11,12,13], there hasbeen a dearth of biophysical studies on the of motion for PIS relative to cholesterol in similar conformation and dynamics of PIS inmembranes
To examine membrane hosts and revealed correlation times close this problem we first employed the techniques of ESR specto thefatty acyl methyl termini in phosphatidylcholine. troscopy of head group nitroxide labeled PIS inmodel mem

Summary

EXPERIMENTAL PROCEDURES

Materials-Geraniol(Clo-OH) and farnesol (Cl,-OH)were purchased from Aldrich. Geraniol was entirely trans isomer; farnesol was a) Dolichol:. For membranes produce a powderpattern in the2H NMR spec- routine work, a 4.5 X 250-mm analytical octadecyl reverse phase trum due to the anisotropic motion of the lipid within the bilayer. For high resolution 'H NMR, the deuterium-la~ledPI ester was dissolved in spectrophotometric grade CHCI, (Aldrich) in 8-mm tubes. 2measurements were formed by a modification a measureof the magnitude of the ~uctuationosf a C-D bond of a composite pulse TI experiment [24] to include the quadrupole vector about its average o ~ e ~ ~ tIn~ goenner.al, a large AVQ echo at the endof the TIpulse sequence for data acquisition. The deuterium spin lattice relaxation time is a sensitive indicator of rates of lipid motions in membranes [19,20,21].

RESULTS

DISCUSSION

AYQmeasurements of PIS are limitedby an inability to TABLEI

In CHCL