Abstract
We here describe a quick and easy method to quantitatively measure in vitro acetylation activity of not only N-terminal acetyltransferase (NAT) enzymes, but acetyltransferases using acetyl-coenzyme A as an acetyl donor in general.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Arnesen T, Van Damme P, Polevoda B, Helsens K, Evjenth R, Colaert N, Varhaug JE, Vandekerckhove J, Lillehaug JR, Sherman F, Gevaert K (2009) Proteomics analyses reveal the evolutionary conservation and divergence of N-terminal acetyltransferases from yeast and humans. Proc Natl Acad Sci USA 106(20):8157–8162. doi:10.1073/pnas.0901931106 pii:0901931106
Aksnes H, Hole K, Arnesen T (2015) Molecular, cellular, and physiological significance of N-terminal acetylation. Int Rev Cell Mol Biol 316:267–305. doi:10.1016/bs.ircmb.2015.01.001
Van Damme P, Hole K, Pimenta-Marques A, Helsens K, Vandekerckhove J, Martinho RG, Gevaert K, Arnesen T (2011) NatF contributes to an evolutionary shift in protein N-terminal acetylation and is important for normal chromosome segregation. PLoS Genet 7(7):e1002169. doi:10.1371/journal.pgen.1002169 pii:PGENETICS-D-11-00127
Aksnes H, Van Damme P, Goris M, Starheim KK, Marie M, Stove SI, Hoel C, Kalvik TV, Hole K, Glomnes N, Furnes C, Ljostveit S, Ziegler M, Niere M, Gevaert K, Arnesen T (2015) An organellar nalpha-acetyltransferase, naa60, acetylates cytosolic N termini of transmembrane proteins and maintains Golgi integrity. Cell Rep 10(8):1362–1374. doi:10.1016/j.celrep.2015.01.053
Dinh TV, Bienvenut WV, Linster E, Feldman-Salit A, Jung VA, Meinnel T, Hell R, Giglione C, Wirtz M (2015) Molecular identification and functional characterization of the first Nalpha-acetyltransferase in plastids by global acetylome profiling. Proteomics 15(14):2426–2435. doi:10.1002/pmic.201500025
Kalvik TV, Arnesen T (2013) Protein N-terminal acetyltransferases in cancer. Oncogene 32(3):269–276. doi:10.1038/onc.2012.82 pii:onc201282
Popp B, Stove SI, Endele S, Myklebust LM, Hoyer J, Sticht H, Azzarello-Burri S, Rauch A, Arnesen T, Reis A (2015) De novo missense mutations in the NAA10 gene cause severe non-syndromic developmental delay in males and females. Eur J Hum Genet 23(5):602–609. doi:10.1038/ejhg.2014.150
Rope AF, Wang K, Evjenth R, Xing J, Johnston JJ, Swensen JJ, Johnson WE, Moore B, Huff CD, Bird LM, Carey JC, Opitz JM, Stevens CA, Jiang T, Schank C, Fain HD, Robison R, Dalley B, Chin S, South ST, Pysher TJ, Jorde LB, Hakonarson H, Lillehaug JR, Biesecker LG, Yandell M, Arnesen T, Lyon GJ (2011) Using VAAST to identify an X-linked disorder resulting in lethality in male infants due to N-terminal acetyltransferase deficiency. Am J Hum Genet 89(1):28–43. doi:10.1016/j.ajhg.2011.05.017 pii:S0002-9297(11)00210-2
Evjenth R, Hole K, Ziegler M, Lillehaug JR (2009) Application of reverse-phase HPLC to quantify oligopeptide acetylation eliminates interference from unspecific acetyl CoA hydrolysis. BMC Proc 3(Suppl 6):S5. doi:10.1186/1753-6561-3-S6-S5 pii:1753-6561-3-S6-S5
Evjenth RH, Van Damme P, Gevaert K, Arnesen T (2013) HPLC-based quantification of in vitro N-terminal acetylation. Methods Mol Biol 981:95–102. doi:10.1007/978-1-62703-305-3_7
Drazic A, Arnesen T (2017) [14C]-Acetyl-coenzyme A-based in vitro N-terminal acetylation assay. In: Schilling O (ed) Protein terminal profiling: methods and protocols, Methods in molecular biology, vol 1574. Springer Science+Business Media LLC, New York, NY
Ellman GL (1959) Tissue sulfhydryl groups. Arch Biochem Biophys 82(1):70–77
Riddles PW, Blakeley RL, Zerner B (1983) Reassessment of Ellman’s reagent. Methods Enzymol 91:49–60
Acknowledgments
These studies were supported by grants from the Norwegian Cancer Society (to T.A.), The Bergen Research Foundation BFS (to T.A.), the Research Council of Norway (grant 230865 to T.A), and the Western Norway Regional Health Authority (to T.A.).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2017 Springer Science+Business Media LLC
About this protocol
Cite this protocol
Foyn, H., Thompson, P.R., Arnesen, T. (2017). DTNB-Based Quantification of In Vitro Enzymatic N-Terminal Acetyltransferase Activity. In: Schilling, O. (eds) Protein Terminal Profiling. Methods in Molecular Biology, vol 1574. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6850-3_2
Download citation
DOI: https://doi.org/10.1007/978-1-4939-6850-3_2
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6849-7
Online ISBN: 978-1-4939-6850-3
eBook Packages: Springer Protocols