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Rowe EM, Biggar KK. An optimized method using peptide arrays for the identification of . MethodsX. 2018;5:118-124doi: 10.1016/j.mex.2018.01.012.
Rowe, E. M., & Biggar, K. K. (2018). An optimized method using peptide arrays for the identification of . MethodsX, 5118-124. https://doi.org/10.1016/j.mex.2018.01.012
Rowe, Elyn M, and Biggar, Kyle K. "An optimized method using peptide arrays for the identification of ." MethodsX vol. 5 (2018): 118-124. doi: https://doi.org/10.1016/j.mex.2018.01.012
Rowe EM, Biggar KK. An optimized method using peptide arrays for the identification of . MethodsX. 2018 Jan 31;5:118-124. doi: 10.1016/j.mex.2018.01.012. eCollection 2018. PMID: 29487803; PMCID: PMC5814365.
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MethodsX. 2018 Jan 31;5:118-124. doi: 10.1016/j.mex.2018.01.012. eCollection 2018.

An optimized method using peptide arrays for the identification of .

MethodsX

Elyn M Rowe, Kyle K Biggar

Affiliations

  1. Institute of Biochemistry and Department of Biology, Carleton University, 1125 Colonel By Drive, Ottawa Ontario, K1N 5B6 Canada.

PMID: 29487803 PMCID: PMC5814365 DOI: 10.1016/j.mex.2018.01.012

Abstract

While a number of post-translational modifications (PTM), such as phosphorylation and ubiquitination, have been extensively studied, lysine methylation is emerging as an important PTM with implications in a growing number of diverse cellular processes. To date, there are approximately 5000 identified methylation sites on non-histone proteins, and as the methyllysine proteome expands it becomes important to identify the lysine methyltransferase enzymes responsible for each methylation event. The use of peptide SPOT methylation assay has proven to be a useful in the identification and validation of novel substrates for lysine methyltransferase enzymes as it uses a weak beta emitter coupled with fluorography to detect methylation events. The method described in this paper provides improvements to the typical protocol for this assay, as a highly sensitive tritium assay can be developed with less radioactivity than previously described. This protocol provides an inexpensive alternative to weak beta signal enhancer sprays and washes for use in lysine methylation peptide SPOT arrays, and a simple open-source method for array quantification.

Keywords: Highly sensitive in vitro SPOT methylation assay; MAP3K2; SMYD3; VEGFR1; non-histone methylation; proteomics

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