Through the process of protein immobilization we can heterogenize enzymes, making it practical to study, use, and combine them in an easily controlled system. In this work, we develop a platform that allows for the simple and oriented immobilization of enzymes through DNA directed immobilization (DDI). Using a previously reported oxidative coupling reaction, we were able to site-selectively conjugate single stranded DNA onto the N-terminus of aldolase from rabbit muscle. This method allows for the generalizability to a wide range of proteins. We were then able to modify the same oxidative coupling chemistry to attach the complementary DNA strand onto a glass surface. Taking advantage of the specificity of Watson-Crick base pairing allowed for the chemically mild, tunable, and reversible method for enzyme immobilization onto glass. Each step of the immobilization process was characterized using fluorescent reporters, atomic force microscopy, and sum frequency generation vibrational spectroscopy. We also conducted activity assays on the immobilized enzymes to determine that the enzyme retained its catalytic activity upon immobilization and were additionally reusable in subsequent cycles.

Site-selective approach for DNA directed immobilization of enzymes via oxidative coupling

253rd National Meeting (2017)

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