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FOR THE RECORD

Designed heterodimerizing leucine zippers with a ranger of pIs and stabilities up to 10^-15 M

¹ Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA
² Laboratory of Biochemistry, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA
³ Laboratory of Molecular Biology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA

Reprint requests to: Charles Vinson, Building 37, Room 4D06, Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA; e-mail: Vinsonc{at}dc37a.nci.nih.gov; fax: (301) 496-8419.

We have designed a heterodimerizing leucine zipper system to target a radionuclide to prelocalized noninternalizing tumor-specific antibodies. The modular nature of the leucine zipper allows us to iteratively use design rules to achieve specific homodimer and heterodimer affinities. We present circular-dichroism thermal denaturation measurements on four pairs of heterodimerizing leucine zippers. These peptides are 47 amino acids long and contain four or five pairs of electrostatically attractive g e' (i, i' +5) interhelical heterodimeric interactions. The most stable heterodimer consists of an acidic leucine zipper and a basic leucine zipper that melt as homodimers in the micro (T_m = 28°C) or nanomolar (T_m = 40°C) range, respectively, but heterodimerize with a T_m >90°C, calculated to represent femtamolar affinities. Modifications to this pair of acidic and basic zippers, designed to destabilize homodimerization, resulted in peptides that are unstructured monomers at 4 µM and 6°C but that heterodimerize with a T_m = 74°C or K_d(37) = 1.1 x 10^-11 M. A third heterodimerizing pair was designed to have a more neutral isoelectric focusing point (pI) and formed a heterodimer with T_m = 73°C. We can tailor this heterodimerizing system to achieve pharmacokinetics aimed at optimizing targeted killing of cancer cells.

Keywords: Leucine zipper; dimerization; electrostatic interactions; radioimmunotherapy; salt bridges; heterodimer

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