The overall objective of this research project is to reveal the chemical basis for the recognition and cleavage of nucleic acids by proteins, and to create novel proteins with properties useful to biology and medicine. The object of this proposal is ribonuclease A (RNase A), a small protein that catalyzes the cleavage of the P-O5 bond of RNA specifically after pyrimidine residues. Classic work on RNase A has contributed much information on enzymatic catalysis; on the chemistry, folding, and stability of proteins; and on molecular evolution. This wealth of information, combined with the ability to produce mutant enzymes and synthesize substrate analogs, make this system ideal for revealing atomic determinants of both enzymatic catalysis and protein - nucleic acid interactions.
The specific aims of this research project are to elucidate (1) the structure of the transition state in RNA cleavage, (2) the mechanism and energetics of enzymatic processivity, (3) the importance of electrostatics in protein - nucleic acid interactions, and (4) how active-site residues cooperate during catalysis. In addition, hybrid proteins will be used to determine whether the divergence of surface loops has allowed RNase.A to acquire special biological activities. Also, semisynthetic ribonucleases will be created that can suffer specific site. Finally, a most important aim is to determine three-dimensional structures of meaningful protein - nucleic acid complexes. The proposed experiments use techniques from organic chemistry, biochemistry, biophysics, and molecular biology to test hypotheses on the relationship between protein structure and protein function (or dysfunction). The results of these experiments will provide new insights into this relationship as well as novel proteins for biomedical analyses and therapies.
| Windsor, Ian W; Palte, Michael J; Lukesh 3rd, John C et al. (2018) Sub-picomolar Inhibition of HIV-1 Protease with a Boronic Acid. J Am Chem Soc 140:14015-14018 |
| Chyan, Wen; Kilgore, Henry R; Raines, Ronald T (2018) Cytosolic Uptake of Large Monofunctionalized Dextrans. Bioconjug Chem 29:1942-1949 |
| Chyan, Wen; Raines, Ronald T (2018) Enzyme-Activated Fluorogenic Probes for Live-Cell and in Vivo Imaging. ACS Chem Biol 13:1810-1823 |
| Chyan, Wen; Kilgore, Henry R; Gold, Brian et al. (2017) Electronic and Steric Optimization of Fluorogenic Probes for Biomolecular Imaging. J Org Chem 82:4297-4304 |
| Mix, Kalie A; Lomax, Jo E; Raines, Ronald T (2017) Cytosolic Delivery of Proteins by Bioreversible Esterification. J Am Chem Soc 139:14396-14398 |
| Smith, Thomas P; Windsor, Ian W; Forest, Katrina T et al. (2017) Stilbene Boronic Acids Form a Covalent Bond with Human Transthyretin and Inhibit Its Aggregation. J Med Chem 60:7820-7834 |
| Hoang, Trish T; Smith, Thomas P; Raines, Ronald T (2017) A Boronic Acid Conjugate of Angiogenin that Shows ROS-Responsive Neuroprotective Activity. Angew Chem Int Ed Engl 56:2619-2622 |
| Burke, Eileen G; Gold, Brian; Hoang, Trish T et al. (2017) Fine-Tuning Strain and Electronic Activation of Strain-Promoted 1,3-Dipolar Cycloadditions with Endocyclic Sulfamates in SNO-OCTs. J Am Chem Soc 139:8029-8037 |
| Newberry, Robert W; Raines, Ronald T (2016) A prevalent intraresidue hydrogen bond stabilizes proteins. Nat Chem Biol 12:1084-1088 |
| Mix, Kalie A; Aronoff, Matthew R; Raines, Ronald T (2016) Diazo Compounds: Versatile Tools for Chemical Biology. ACS Chem Biol 11:3233-3244 |
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