Folding and Self-Association Studies of Id3 Dimerization Domain

The Id3 protein is a member of the family of the Id proteins (Id1-4), which act as inhibitors of DNA binding and cell differentiation and represent the class V of the large family of the helix-loop-helix (HLH) transcription factors. The Id proteins are potential targets in cancer therapy, as they are upregulated in several types of tumors, promoting tumor growth and metastasis. Besides heterodimerization with basic-HLH transcription factors, the Id3 protein can also undergo self-association, which is driven by the HLH domain. Three different approaches to study and control the self-association of the Id3 HLH domain have been applied in this work. One approach is based on the use of thiol-containing analogs and oxidation/disulfide reshuffling assays, the second one is based on electrostatic interactions between a positively charged tag of an Id3 HLH analog and a negatively charged one of another analog, and the last approach is based on the use of fluorescent peptide analogs to be investigated by FRET spectroscopy.