Date of Graduation


Document Type


Degree Name

Doctor of Philosophy in Chemistry (PhD)

Degree Level



Chemistry & Biochemistry


Roger E. Koeppe

Committee Member

Dan Davis

Second Committee Member

Paul Adams

Third Committee Member

David Paul

Fourth Committee Member

Suresh Kumar Krishnaswamy Thallapuranam


Pure sciences, Biological sciences, Aromatic anchoring residues, Gala, Lipid, Membranes, Protein-lipid interactions, Transmembrane, Walp


To investigate in detail the interactions between transmembrane proteins and the lipid bilayers in which they are constituted, designed model peptides with selective isotopic labels were synthesized and analyzed by means of solid-state deuterium NMR spectroscopy. Starting from the well-characterized model peptide GWALP23, acetyl-GGALW(LA)6LWLAGA-amide, several Trp to Tyr mutations were compared to evaluate their respective interfacial anchoring abilities. It was found that Tyr, substituted on either or both termini, can effectively anchor the transmembrane alpha-helix, which then adopts a similar transmembrane topology in a range of bilayer thicknesses. Nevertheless, a consistent ~10° shift in tilt direction (helix rotation) is observed when a Tyr is substituted for Trp and found to be terminal-dependent (i.e. in opposite direction on each end). The fluorescence emission spectra from the single remaining Trp residue in Y5GWALP23 and Y19GWALP23 indicate that W5 is buried more deeply in the bilayer than is W19.

Using Y5GWALP23 as a host, the influence of Lys was examined at positions 12 and 14 in various lipid bilayer thicknesses. Y5GWALP23-K14 incorporates well into both thin and thick bilayers. It influences the peptide's orientation by increasing the tilt magnitude (4-9°) and altering the tilt direction (60-95°). In contrast, the L12K mutant yields multiple low-intensity peaks in 2H NMR spectra, recorded in DOPC, indicative of multi-state behavior. Nevertheless, the peptide orients well and adopts a large tilt angle (30°) in the thinner bilayers of DLPC. Y5GWALP23-K12 in DOPC is observed to titrate at high pH to a neutral form that is well aligned in an orientation that is very similar to that of the host peptide without lysine. Titration of Y5GWALP23-K14 reveals a pKa of 6.2 in DOPC at 50 °C and different transmembrane orientations when the peptides charged lysine, neutral lysine or no lysine are compared.

When Glu is added adjacent to a lysine, significantly improved NMR spectra and a stable transmembrane orientation are observed for Y5GWALP23-(K12, E13). Y5GWALP23-K14 experiences a small change in orientation with Glu-15 addition, but interestingly titrates much like the peptide with K14 alone. Placement of a Glu residue (E13) near R12 also improves spectra quality, especially at higher pH. It appears a stabilizing ion-pair may in some instances "rescue" the K12 or R12 peptide from its multi-state behavior in DOPC. The individual ionization states of paired ionizable residues have yet to be determined.