United-atom force fields for molecular dynamics (MD) simulations provide a higher computational efficiency, especially in lipid membrane simulations, with little sacrifice in accuracy, when compared to all-atom force fields. Excellent united-atom lipid models are available, but in combination with depreciated protein force fields. In this work, a united-atom model of the lipid 1,2-dipalmitoyl-sn-glycero-3-phosphocholine has been built with standard parameters of the force field GROMOS96 53a6 that reproduces the experimental area per lipid of a lipid bilayer within 3% accuracy to a value of 0.623 ( 0.011 nm 2 without the assumption of a constant surface area or the inclusion of surface pressure. In addition, the lateral self-diffusion constant and deuterium order parameters of the acyl chains are in agreement with experimental data. Furthermore, models for 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), 1-palmitoyl-2oleoyl-sn-glycero-3-phosphocholine (POPC), and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) result in areas per lipid of 0.625 nm 2 (DMPC), 0.693 nm 2 (POPC), and 0.700 nm 2 (POPG) from 40 ns MD simulations. Experimental lateral self-diffusion coefficients are reproduced satisfactorily by the simulation. The lipid models can form the basis for molecular dynamics simulations of membrane proteins with current and future versions of united-atom protein force fields.

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The coronavirus responsible for the severe acute respiratory syndrome contains a small envelope protein, E, with putative involvement in host apoptosis and virus morphogenesis. To perform these functions, it has been suggested that protein E can form a membrane destabilizing transmembrane (TM) hairpin, or homooligomerize to form a TM pore. Indeed, in a recent study we reported that the alpha-helical putative transmembrane domain of E protein (ETM) forms several SDS-resistant TM interactions: a dimer, a trimer, and two pentameric forms. Further, these interactions were found to be evolutionarily conserved. Herein, we have studied multiple isotopically labeled ETM peptides reconstituted in model lipid bilayers, using the orientational parameters derived from infrared dichroic data. We show that the topology of ETM is consistent with a regular TM alpha-helix. Further, the orientational parameters obtained unequivocally correspond to a homopentameric model, by comparison with previous predictions. We have independently confirmed that the full polypeptide of E protein can also aggregate as pentamers after expression in Escherichia coli. This interaction must be stabilized, at least partially, at the TM domain. The model we report for this pentameric alpha-helical bundle may explain some of the permabilizing properties of protein E, and should be the basis of mutagenesis efforts in future functional studies.

The recently developed method of site-directed Fourier transform infrared dichroism for obtaining orientational constraints of oriented polymers is applied here to the transmembrane domain of the vpu protein from the human immunodeficiency virus type 1 (HIV-1). The infrared spectra of the 31-residue-long vpu peptide reconstituted in lipid vesicles reveal a predominantly alpha-helical structure. The infrared dichroism data of the (13)C-labeled peptide yielded a helix tilt beta = (6.5 +/- 1.7) degrees from the membrane normal. The rotational pitch angle omega, defined as zero for a residue located in the direction of the helix tilt, is omega = (283 +/- 11) degrees for the (13)C labels Val(13)/Val(20) and omega = (23 +/- 11) degrees for the (13)C labels Ala(14)/Val(21). A global molecular dynamics search protocol restraining the helix tilt to the experimental value was performed for oligomers of four, five, and six subunits. From 288 structures for each oligomer, a left-handed pentameric coiled coil was obtained, which best fits the experimental data. The structure reveals a pore occluded by Trp residues at the intracellular end of the transmembrane domain.

17 O solid-state NMR from 14 amino acids is reported here, greatly increasing the number investigated. In most cases well-separated resonances from carbonyl and hydroxyl oxygens with distinct second-order quadrupolar line shapes are observed using a 600 MHz spectrometer with fast magic angle spinning (MAS). This is in contrast to the motionally averaged resonances usually seen from amino acids in solution. For amino acids double-angle rotation (DOR) produces a decrease in the line width by more than a factor of 40, providing very high resolution, ∼1 ppm, spectra. The oxygen lines in alanine and the carbonyl oxygens in L-glutamic acid hydrochloride are assigned using 1 H-decoupled DOR. The NMR interaction parameters for amino acids show a wide variation of Q , from 6.4 to 8.6 MHz, η, from 0.0 to 0.9, and δ iso , from 83 to 353 ppm. The high quality of the MAS NMR line shapes obtained at 14.1 T means that even small changes in parameters can be very accurately deduced, offering the possibility of 17 O NMR as a sensitive probe of structural changes in these and related compounds. The D-and L-forms of glutamic acid hydrochloride are shown to have the same NMR parameters to within error, which are very different from those reported in the literature for the D,L-form. A strong correlation (∼-1200 ppm/Å) is found between δ iso and the C-O bond length of the carbonyl oxygens. On the basis of these data, enriching specific amino acids in more complex polypeptides and proteins could provide site-selective information about the bonding and functionality of different sites in biomolecules. An estimate is made of the possible detection limit for such species.