Proteins carry out many different tasks that are necessary for life. The specific task performed depends on the 3-dimensional structure of the protein molecule. In nature, a protein's structure can be purposely altered through interaction with ions, like calcium, or by chemical modifications, such as adding a sugar group. Even though it is predicted that most proteins have sugars attached to them, forming so-called glycoproteins, only a small proportion of the 100,000+ protein structures studied by scientists are glycoproteins. To understand how attached sugars alter the structure of a calcium-binding protein, the effects of the attached sugars must be systematically investigated. Understanding the synergistic effects of the sugar, calcium, and protein will allow investigators to study the function of these unique protein shapes and to predict the structure of similar glycoproteins. Graduate and undergraduate students from underrepresented groups will be recruited for this project. They will acquire specialized training in protein chemistry, nuclear magnetic resonance spectroscopy, and mass spectrometry.
With this award, NSF-EPSCoR and the Chemistry of Life Processes Program in the Chemistry Division are funding Dr. Megan Macnaughtan from Louisiana State University to determine the synergistic effects of calcium binding and glycosylation on the structure of the mouse Notch1 Abruptex region. The structure and function of epidermal growth factor-like (EGF) repeats, like those in the Notch1 Abruptex region, are regulated by highly conserved glycosylation and calcium-binding sites. The effects of high-affinity calcium-binding sites on the structure of tandem EGF repeats has been well-studied, but the synergistic and singular effects of glycosylation have not been systematically investigated. EGF repeats 26-28 of the Notch1 Abruptex region will be studied to determine the calcium dissociation constants and the structural changes upon sequential glycosylation using fluorescence spectrophotometry and nuclear magnetic resonance (NMR) spectroscopy. The high-resolution 3-dimensional structure of calcium-bound, glycosylated EGF26-28 will be determined using NMR to identify important interactions. By defining the interactions between calcium, the glycans, and the protein, new insights into the structure of other glycosylated EGF repeats and the regulation of EGF repeat function will be revealed, impacting the fields of structural biology and cellular signaling.
