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Peptide Microarrays to Examine RNA Polymerase II Binding Protein Domains
Author(s) -
Lothrop Adam,
Can Joe,
Fuchs Stephen
Publication year - 2015
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.29.1_supplement.877.12
Subject(s) - ctd , rna polymerase ii , biology , rna binding protein , transcription (linguistics) , computational biology , protein subunit , c terminus , microbiology and biotechnology , rna , biochemistry , gene , promoter , amino acid , gene expression , oceanography , linguistics , philosophy , geology
RNA polymerase II (RNAP II) is responsible for the majority of mRNA transcription in eukaryotes. The C‐terminal domain (CTD) of the largest RNAP II subunit is repetitive in amino acid sequence and extensively post‐translationally modified. The CTD forms a scaffold for transcriptionally‐associated factors that bind the polymerase, with temporal regulation of binding mediated by patterns of post‐translational modifications (PTMs). In order to ascertain how combinatorial modifications affect binding of these factors to the CTD, we have created a library of CTD peptides with specific combinations of PTMs. The library is used in a high‐throughput peptide microarray approach in conjunction with additional biophysical techniques to study binding of CTD associated proteins. Here, we have focused on domains of several serine‐arginine rich (SR) proteins that share low sequence identity, but high structural homology to the CTD binding domain of the methyltransferase SetD2. Use of the combinatorially modified library enables us to examine how different PTMs affect CTD binding by these SR‐proteins, many of which are implicated in human disease. Supported by NIH post‐doctoral fellowship F32 GM106588‐02 to A.P.L.