PREFERENTIAL HETERODIMERIC PARALLEL COILED-COIL FORMATION BY SYNTHETIC MAX AND C-MYC LEUCINE ZIPPERS - A DESCRIPTION OF PUTATIVE ELECTROSTATIC INTERACTIONS RESPONSIBLE FOR THE SPECIFICITY OF HETERODIMERIZATION

The oncoprotein c-Myc must heterodimerize with Max to bind DNA and per form its oncogenic activity. The c-Myc-Max heterodimer binds DNA throu gh a basic helix-loop-helix leucine zipper (b-HLH-zip) motif and it is proposed that leucine zipper domains could, in concert with the HLH r egions, provide the specificity and stability of the b-HLH-zip motif. In this context, we have synthesized the peptides corresponding to the leucine zipper domains of Max and c-Myc with a N-terminal Cys-Gly-Gly linker and studied their dimerization behavior using reversed-phase H PLC and CD spectroscopy. The preferential formation of a fully helical parallel c-Myc-Max heterodimeric coiled-coil was observed under air-o xidation and redox conditions at neutral pH. We show that the stabilit y and the helicity of the disulfide-linked c-Myc-Max heterostranded co iled-coil is modulated by pH, with a maximum around pH 4.5, supporting the existence of stabilizing and specific interhelical electrostatic interactions. We present a molecular model of the c-Myc-Max heterostra nded coiled-coil describing potential electrostatic interactions respo nsible for the specificity of the interaction, the main feature being putative buried electrostatic interactions between a histidine side-ch ain (in the Max leucine zipper) and two glutamic acid side-chains (in the c-Myc leucine zipper) at the heterodimer interface. This model is supported by the fact that the apparent pKa (as determined by [H-1]-NM R spectroscopy) of this histidine side-chain at 25 degrees C is 0.42 ( +/-0.05) pKa units higher in the folded form than in the unfolded form . This indicates that the charged histidine side-chain contributes app roximately 0.57 (+/-0.07) kcal/mol (2.38 (+/-0.30) kJ/mol) of stabiliz ation free energy to the c-Myc-Max heterostranded coiled-coil through favorable electrostatic interaction. (C) 1995 Academic Press Limited