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Monitoring AMP production in diverse biochemical reactions using HTS‐formatted bioluminescent assays
Author(s) -
Goueli Said,
Hsiao Kevin,
Zegzouti Hicham
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.663.14
Subject(s) - enzyme , ubiquitin , biochemistry , dna , dna repair , phosphodiesterase , biology , nucleotide , chemistry , gene
Adenine nucleotides are major determinants of the energy status of the cell and thus any modulation of their cellular concentration has significant consequences to cellular metabolism, cellular growth and cell death. Many biochemical reactions that generate AMP as a reaction product are pivotal players in a variety of signaling pathways and thus are considered validated drug targets. These include enzymes involved in protein ubiquitination, aminoacyl tRNA synthetases in protein synthesis, cAMP‐Phosphodiesterases in second messenger regulation, DNA ligases in DNA synthesis and DNA repair, etc. Thus an assay that can monitor the concentration of AMP generated by these enzymes is desirable in the search for modulators of these enzymes and the development of novel therapeutics. In addition, enzyme reactions that generate products which can be converted to AMP can be also monitored using this assay. We have developed a homogenous bioluminescent assay that is antibody free, sensitive, free from fluorescent compound interference, and easy to use. We will show its utility in various applications such as cAMP‐phosphodiesterases, ubiquitin ligases, DNA ligases, methyltransferases, etc