Abstract
Here we describe the procedure for expression and purification of recombinant ATE1 from E. coli. This method is easy and convenient and can result in one-step isolation of milligram amounts of soluble enzymatically active ATE1 at nearly 99 % purity. We also describe a procedure for expression and purification of E. coli Arg-tRNA synthetase essential for the arginylation assays described in the next two chapters.
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Saha S, Wang J, Buckley B, Wang Q, Lilly B, Chernov M, Kashina A (2012) Small molecule inhibitors of arginyltransferase regulate arginylation-dependent protein degradation, cell motility, and angiogenesis. Biochem Pharmacol 83(7):866–873
Wang J, Han X, Saha S, Xu T, Rai R, Zhang F, Wolf YI, Wolfson A, Yates JR III, Kashina A (2011) Arginyltransferase is an ATP-independent self-regulating enzyme that forms distinct functional complexes in vivo. Chem Biol 18(1):121–130
Wang J, Han X, Wong CC, Cheng H, Aslanian A, Xu T, Leavis P, Roder H, Hedstrom L, Yates JR III, Kashina A (2014) Arginyltransferase ATE1 catalyzes midchain arginylation of proteins at side chain carboxylates in vivo. Chem Biol 21(3):331–337
Rai R, Kashina A (2005) Identification of mammalian arginyltransferases that modify a specific subset of protein substrates. Proc Natl Acad Sci U S A 102(29):10123–10128
Rai R, Mushegian A, Makarova K, Kashina A (2006) Molecular dissection of arginyltransferases guided by similarity to bacterial peptidoglycan synthases. EMBO Rep 7(8):800–805
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© 2015 Springer Science+Business Media New York
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Wang, J., Kashina, A.S. (2015). Bacterial Expression and Purification of Recombinant Arginyltransferase (ATE1) and Arg-tRNA Synthetase (RRS) for Arginylation Assays. In: Kashina, A. (eds) Protein Arginylation. Methods in Molecular Biology, vol 1337. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2935-1_9
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DOI: https://doi.org/10.1007/978-1-4939-2935-1_9
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2934-4
Online ISBN: 978-1-4939-2935-1
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