The dimerization stability of the HLH-LZ transcription protein family is modulated by the leucine zippers: a CD and NMR study of TFEB and c-Myc.

In the HLH-LZ protein family, the helix-loop-helix DNA-binding dimerization domain is followed in the sequence by a leucine zipper motif. The precise function of this second dimerization domain is still unclear, since the HLH motif of a subset of this family has been shown to be necessary and sufficient for dimerization. However, deletion and mutagenesis studies of the leucine zipper in various HLH-LZ proteins have shown a clear influence of this motif on homo- and heterodimerization. In this paper, we present a structural characterization of synthetic peptides encompassing the leucine zipper sequences of c-Myc and TFEB, using circular dichroism, analytical ultracentrifugation, and nuclear magnetic resonance. We show that the different ability of the synthetic leucine zippers of c-Myc and TFEB to homodimerize at neutral pH reflects the different dimerization properties reported for the entire proteins. The TFEB protein is known to form homodimers. c-Myc, on the other hand, does not homodimerize in vivo, but is mostly found in heterodimeric complexes with Max, another protein of the HLH-LZ family. Accordingly, our results show that the TFEB peptide homodimerizes at neutral pH whereas the Myc peptide dimerizes to a comparable amount only at acidic pH and high ionic strength. Both synthetic peptides are far less stable than leucine zippers of the b-ZIP family. The relative stability of the two leucine zippers and the factors which stabilize the dimer formation are discussed.