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Nucleic Acids Research, 2001, Vol. 29, No. 2 479-487
© 2001 Oxford University Press

DNA-bound transcription factor complexes analysed by mass-spectrometry: binding of novel proteins to the human c-fos SRE and related sequences

Victoria Drewett, Henrik Molina1, Alan Millar2, Silke Muller, Friedrich von Hesler and Peter E. Shaw*

School of Biomedical Sciences, D Floor, University of Nottingham Medical School, Queen’s Medical Centre, Nottingham NG7 2UH, UK, 1Protana, Staermosegaardsvej 16, DK-5230 Odense M, Denmark and 2micromass UK, Floats Road, Wythenshawe, Manchester M23 9LZ, UK

Transcription factors control eukaryotic polymerase II function by influencing the recruitment of multiprotein complexes to promoters and their subsequent integrated function. The complexity of the functional ‘transcriptosome’ has necessitated biochemical fractionation and subsequent protein sequencing on a grand scale to identify individual components. As a consequence, much is now known of the basal transcription complex. In contrast, less is known about the complexes formed at distal promoter elements. The c-fos SRE, for example, is known to bind Serum Response Factor (SRF) and ternary complex factors such as Elk-1. Their interaction with other factors at the SRE is implied but, to date, none have been identified. Here we describe the use of mass-spectrometric sequencing to identify six proteins, SRF, Elk-1 and four novel proteins, captured on SRE duplexes linked to magnetic beads. This approach is generally applicable to the characterisation of nucleic acid-bound protein complexes and the post-translational modification of their components.

* To whom correspondence should be addressed. Tel: +44 115 970 9362; Fax: +44 115 970 9926; Email: peter.shaw{at}nottingham.ac.uk


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