A novel approach for studying biological proton transfer: Protein incorporation...
A novel approach for studying biological proton transfer: Protein incorporation of noncanonical amino acids carrying a light-triggered proton donor and proton acceptor
Proton transfer (PT) reactions within proteins are fundamental in biological systems, such as within ATP production. To date, there are no direct means to measure specific PT pathways within proteins, and most research is based on...
Proton transfer (PT) reactions within proteins are fundamental in biological systems, such as within ATP production. To date, there are no direct means to measure specific PT pathways within proteins, and most research is based on following the end-product of the PT reaction across a natural proton pathway. Here, our goal is to develop a novel way to directly measure PT within proteins. Our new approach is based on placing proton donors and acceptors in specific places within proteins, in which the PT will be initiated only after light excitation of the system. To do so, we introduce here two noncanonical amino acids (ncAA) we developed, with a photoacid and a photobase as their residues that serve as the proton donor and acceptor, respectively. Our hypothesis is that the strong light-triggered driving force of PT in the excited-state (of ~11 pKa units) will initiate PT along the pathway from donor to acceptor. In the first objective, we will use our new ncAA with solid-phase peptide synthesis to design several peptide systems that will allow us to decipher the role of specific amino acids, the peptide structure, and the role of water in PT across the peptide using various ultrafast spectroscopy. In the second objective, we aim to design a mutually orthogonal system for the insertion of our two ncAA into a single protein. In our third objective, we plan to use our new experimental system for answering a unique set of questions in the field of biological PT that could not have been answered before, focusing on the systems of ATP synthase transmembrane complex and the soluble carbonic anhydrase enzyme. Our new approach is groundbreaking in the way we study and understand PT in biology and will enable researchers completely new capabilities resulting in fascinating new discoveries. Moreover, our new system can be translated into other fields in biology that require the local change in proton concentration within proteins.ver más
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