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Novel subunits of the TATA binding protein free TAF II ‐containing transcription complex identified by matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry following one‐dimensional gel electrophoresis
Author(s) -
Cavusoglu Nükhet,
Brand Marjorie,
Tora Laszlo,
Van Dorsselaer Alain
Publication year - 2003
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200390030
Subject(s) - chemistry , mass spectrometry , tata binding protein , gel electrophoresis , rna polymerase ii , microbiology and biotechnology , transcription factor ii d , rna polymerase iii , transcription factor , biology , rna , dna binding protein , rna polymerase , biochemistry , chromatography , promoter , gene expression , gene
Initiation of transcription of protein‐encoding genes by RNA polymerase II was thought to require the transcription factor II D (TF II D), a complex comprising the TATA binding protein (TBP) and TBP‐associated factors. However, another multiprotein complex isolated more recently and called TFTC (TBP‐free TAF II containing complex), was shown to mediate initiation of RNA polymerase II (Pol II) transcription in the absence of TF II D as well as specific acetylation of histone H3 in a nucleosomal context. Several subunits of the TFTC complex were already identified using classical methods such as Edman based microsequencing and Western blot analysis. In this article we present a mass spectrometry based proteomic approach to confirm previous results and to identify other possible subunits of the TFTC complex. The TFTC complex was separated on one‐dimensional sodium dodecyl sulfate polyacrylamide electrophoresis and analysed by matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry and peptide mass fingerprinting. Identifications were realized after databank searches. This new characterization of TFTC complex confirmed the presence of already described subunits (TRRAP, GCN5, SAP130/KIA0017, TAF II 150, TAF II 135, TAF II 100, TAF II 80, TAF II 20, SPT3 and PAF65β). Moreover, a good coverage of these sequences was obtained. Interestingly, TAF II 32 and PAF6α were also determined as potential novel subunits of TFTC. These results together show the suitability and the great potential of this method and offer new perspectives in fundamental studies of transcription factor complexes.