RESUMEN
Introduction: Pilot projects ("pilots") are important for testing hypotheses in advance of investing more funds for full research studies. For some programs, such as Clinical and Translational Science Awards (CTSAs) supported by the National Center for Translational Sciences, pilots also make up a significant proportion of the research projects conducted with direct CTSA support. Unfortunately, administrative data on pilots are not typically captured in accessible databases. Though data on pilots are included in Research Performance Progress Reports, it is often difficult to extract, especially for large programs like the CTSAs where more than 600 pilots may be reported across all awardees annually. Data extraction challenges preclude analyses that could provide valuable information about pilots to researchers and administrators. Methods: To address those challenges, we describe a script that partially automates extraction of pilot data from CTSA research progress reports. After extraction of the pilot data, we use an established machine learning (ML) model to determine the scientific content of pilots for subsequent analysis. Analysis of ML-assigned scientific categories reveals the scientific diversity of the CTSA pilot portfolio and relationships among individual pilots and institutions. Results: The CTSA pilots are widely distributed across a number of scientific areas. Content analysis identifies similar projects and the degree of overlap for scientific interests among hubs. Conclusion: Our results demonstrate that pilot data remain challenging to extract but can provide useful information for communicating with stakeholders, administering pilot portfolios, and facilitating collaboration among researchers and hubs.
RESUMEN
Parallel ß-sheet-containing repeat proteins often display a structural motif in which conserved asparagines form a continuous ladder buried within the hydrophobic core. In such "asparagine ladders", the asparagine side-chain amides form a repetitive pattern of hydrogen bonds with neighboring main-chain NH and CO groups. Although asparagine ladders have been thought to be important for stability, there is little experimental evidence to support such speculation. Here we test the contribution of a minimal asparagine ladder from the leucine-rich repeat protein pp32 to stability and investigate lattice rigidity and hydrogen bond character using solution nuclear magnetic resonance (NMR) spectroscopy. Point substitutions of the two ladder asparagines of pp32 are strongly destabilizing and decrease the cooperativity of unfolding. The chemical shifts of the ladder side-chain HZ protons are shifted significantly downfield in the NMR spectrum and have low temperature coefficients, indicative of strong hydrogen bonding. In contrast, the HE protons are shifted upfield and have temperature coefficients close to zero, suggesting an asymmetry in hydrogen bond strength along the ladder. Ladder NH2 groups have weak 1H-15N cross-peak intensities; 1H-15N nuclear Overhauser effect and 15N CPMG experiments show this to be the result of high rigidity. Hydrogen exchange measurements demonstrate that the ladder NH2 groups exchange very slowly, with rates approaching the global exchange limit. Overall, these results show that the asparagine side chains are held in a very rigid, nondynamic structure, making a significant contribution to the overall stability. In this regard, buried asparagine ladders can be considered "second backbones" within the cores of their elongated ß-sheet host proteins.
