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- Title
Use of a Two-Hybrid System to Identify Mutations in Max That Confer Increased Affinity for Myc.
- Authors
WANG, HONGBING; VAN DEN BERGH, FRANCOISE; SPENCER, ERIKA; WILCOX, KENT; HERMAN, TIM
- Abstract
A yeast two-hybrid system was used to identify mutants of Max that exhibit an increased affinity for Myc. Truncated forms of the Max helix-loop-helix/leucine zipper motif (HLH/Zip) were first expressed in a twohybrid system in which the bait protein was the HLH/Zip motif of Myc. Deletion of amino acids both aminoterminal and carboxy-terminal to the leucine zipper of Max reduced Myc/Max heterodimer formation as evidenced by a 160-fold reduction in the expression of the lacZ gene. A library of partially randomized sequences encoding this minimal leucine zipper of Max was then screened using the two-hybrid system. Mutant forms of the Max leucine zipper were identified whose affinities for Myc, as measured by β-galactosidase activity in yeast lysates, were from 8- to 200-fold greater than the wild-type Max zipper. These Max mutants were shown to interact specifically with Myc and not with wild-type Max. Of 29 mutants analyzed, all had a unique amino acid sequence. This result illustrates the value of a genetic screen in the identification of a collection of mutant forms of the Max leucine zipper whose structures would not have been predicted based on principles of structure-based design.
- Publication
DNA & Cell Biology, 1997, Vol 16, Issue 11, p1277
- ISSN
1044-5498
- Publication type
Article
- DOI
10.1089/dna.1997.16.1277