Selection of abstracts from Baylor Scott & White Health Central Texas Scholars Day.

Baylor Scott & White Health Central Texas displayed the diversity and growth of scholarly pursuits during Scholars Day on May 3, 2019. Residents and fellows, medical students, nurses, and research staff were among those showcasing their scholarly activity in areas such as medical innovation, clinical vignettes, research, and quality improvement. A selection committee chose 32 abstracts-12 select podium, 20 rapid fire. In addition, 60 abstracts were included as electronic poster presentations. Residency and fellowship program directors nominated presentations for the Excellence Awards. The scholarship committee chose four to receive travel awards to support the presentation of each project at a national meeting. Excellence Awards were granted to Jasson Abraham, MD, Jerry Fan, MD, Veronica Lozano, MD, and Chhaya Patel, MD. A selection of abstracts is presented here.

Detailed microscopic unfolding pathways of an α-helix and a ß-hairpin: direct observation and molecular dynamics.

We present a combined experimental and computational study of unfolding pathways of a model 21-residue α-helical heteropeptide (W1H5-21) and a 16-residue ß-hairpin (GB41-56). Experimentally, we measured fluorescence energy transfer efficiency as a function of temperature, employing natural tryptophans as donors and dansylated lysines as acceptors. Secondary structural analysis was performed with circular dichroism and Fourier transform infrared spectroscopy. Our studies present markedly different unfolding pathways of the two elementary secondary structural elements. During thermal denaturation, the helical peptide exhibits an initial decrease in length, followed by an increase, while the hairpin undergoes a systematic increase in length. In the complementary computational part of the project, we performed microsecond length replica-exchange molecular dynamics simulations of the peptides in explicit solvent, yielding a detailed microscopic picture of the unfolding processes. For the α-helical peptide, we found a large heterogeneous population of intermediates that are primarily frayed single helices or helix-turn-helix motifs. Unfolding starts at the termini and proceeds through a stable helical region in the interior of the peptide but shifted off-center toward the C-terminus. The simulations explain the experimentally observed non-monotonic variation of helix length with temperature as due primarily to the presence of frayed-end single-helix intermediate structures. For the ß-hairpin peptide, our simulations indicate that folding is initiated at the turn, followed by formation of the hairpin in zipper-like fashion, with Cα···Cα contacts propagating from the turn to termini and hairpin hydrogen bonds forming in parallel with these contacts. In the early stages of hairpin formation, the hydrophobic side-chain contacts are only partly populated. Intermediate structures with low numbers of ß-hairpin hydrogen bonds have very low populations. This is in accord with the "broken zipper" model of Scheraga. The monotonic increase in length with temperature may be explained by the zipper-like breaking of the hairpin hydrogen bonds and backbone contacts.

Experiments and comprehensive simulations of the formation of a helical turn.

We investigate the kinetics and thermodynamics of a helical turn formation in the peptide Ac-WAAAH-NH(2). NMR measurements indicate that this peptide has significant tendency to form a structure of a helical turn, while temperature dependent CD establishes the helix fraction at different temperatures. Molecular dynamics and milestoning simulations agree with experimental observables and suggest an atomically detailed picture for the turn formation. Using a network representation, two alternative mechanisms of folding are identified: (i) a direct co-operative mechanism from the unfolded to the folded state without intermediate formation of hydrogen bonds and (ii) an indirect mechanism with structural intermediates with two residues in a helical conformation. This picture is consistent with kinetic measurements that reveal two experimental time scales of sub-nanosecond and several nanoseconds.

Unassisted transport of N-acetyl-L-tryptophanamide through membrane: experiment and simulation of kinetics.

Cellular transport machinery, such as channels and pumps, is working against the background of unassisted material transport through membranes. The permeation of a blocked tryptophan through a 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) membrane is investigated to probe unassisted or physical transport. The transport rate is measured experimentally and modeled computationally. The time scale measured by parallel artificial membrane permeation assay (PAMPA) experiments is ~8 h. Simulations with the milestoning algorithm suggest mean first passage time (MFPT) of ~4 h and the presence of a large barrier at the center of the bilayer. A similar calculation with the solubility-diffusion model yields a MFPT of ~15 min. This permeation rate is 9 orders of magnitude slower than the permeation rate of only a tryptophan side chain (computed by us and others). This difference suggests critical dependence of transport time on permeant size and hydrophilicity. Analysis of the simulation results suggests that the permeant partially preserves hydrogen bonding of the peptide backbone to water and lipid molecules even when it is moving closer to the bilayer center. As a consequence, defects of the membrane structure are developed to assist permeation.

Structural dynamics of neuropeptide hPYY.

We have employed a combination of experiment and simulation to characterize the ensemble of structures sampled by human Peptide YY (hPYY), an important member of the neuropeptide Y family. Experimental structural characterization carried out with far UV circular dichroism spectroscopy and Fourier Transform-Infrared measurements confirmed that the major feature of the secondary structure of hPYY is the α-helix, encompassing about half the peptide residues, with smaller contributions from turn and ß-sheet like structures. The peptide undergoes thermal denaturation characterized by a melting temperature of 48°C with an enthalpy change of -24.5 kcal/mol and entropy change of -76.2 cal/(mol K). In our computational studies, based on a 4-µsec MD trajectory generated with the AMBER03 potential, we found excellent agreement of the predicted features with experimental data, including a stable C-terminal helix, a central turn and conservation of about 80% of measured long-range NOE contacts. The main structural fluctuations involved partial helix unwinding and large-scale motions of the N-terminal. Our joint experimental/computational approach leads to several insights into the biological function of PYY. We conclude that the C-terminal helix is crucial for the structural integrity of PYY. The structures and motions found in the simulations suggest microscopic explanations for observed changes in biological activity of the peptide upon mutation and truncation. We also performed microsecond-length MD and replica-exchange simulations of hPYY with the OPLS-AA force field, for which computed structures did not agree well with experimental data, predicting significant loss of helicity and NOE contacts.

Helix formation in a pentapeptide: experiment and force-field dependent dynamics.

We used a combined approach of experiment and simulation to determine the helical population and folding pathway of a small helix forming blocked pentapeptide, Ac-(Ala)(5)-NH(2). Experimental structural characterization of this blocked peptide was carried out with far UV circular dichroism spectroscopy, FTIR, and NMR measurements. These measurements confirm the presence of the α-helical state in a buffer solution. Direct molecular dynamics and replica-exchange simulations of the pentapeptide were performed using several popular force fields with explicit solvent. The simulations yielded statistically reliable estimates of helix populations, melting curves, folding, and nucleation times. The distributions of conformer populations are used to measure folding cooperativity. Finally, a statistical analysis of the sample of helix-coil transition paths was performed. The details of the calculated helix populations, folding kinetics and pathways vary with the employed force field. Interestingly, the helix populations, folding, and unfolding times obtained from most of the studied force fields are in qualitative agreement with each other and with available experimental data, with the deviations corresponding to several kcal/mol in energy at 300 K. Most of the force fields also predict qualitatively similar transition paths, with unfolding initiated at the C-terminus. Accuracy of potential energy parameters, rather than conformational sampling may be the limiting factor in current molecular simulations.