Reprogramming Escherichia coli with synthetic protein glycosylation pathways

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1159581/ DeLisaAsparagine-linked (N-linked) protein glycosylation is the most common modification made to proteins in eukaryotic cells. N-linked glycosylation pathways are also known to exist in gram-negative bacteria. The bacterial glycosylation system bears many functional similarities with the eukaryotic process, but there are also several major differences between the systems. Most notably, the structures of bacterial N-linked glycans are significantly different from those produced in eukaryotes. Hence, glycoproteins derived from current bacterial glycosylation systems are likely to be immunogenic in humans and thus of limited therapeutic value. This project seeks to take bacterial glycosylation to a previously unexplored level by bottom-up engineering of a synthetic pathway for performing eukaryotic N-linked glycosylation reactions in Escherichia coli. The proposed studies and research training activities are expected to have a broad impact on society, ranging from the science of glycobiology to the development of human glycotherapeutics. This project will also cultivate the next generation of highly trained graduate students who will be introduced to the broad, interdisciplinary nature of biotechnology research. Moreover, this program will actively and aggressively broaden participation in science and engineering. This will be accomplished by collaborating with Cornell's Diversity Programs in Engineering and with the Cornell Louis Stokes Alliance for Minority Participation (LSAMP) Program to develop a hands-on learning experience entitled "How Sweet It Is" that will introduce undergraduate students from underrepresented groups to molecular biotechnology and the important role played by sugars in living systems. Finally, the development of bacterial glycosylation for biotechnological applications will be brought to a larger research community through partnership with local biotechnology companies. Overall, these studies will provide considerable fuel for new academic pursuits and industrial enterprises related to bacterial glycoengineering.