Inactivation of the N-myc gene product by single amino acid substitution of leucine residues located in the leucine-zipper region.
To verify the importance of the hypothetical leucine-zipper structure in the N-myc protein, a series of mutants of the mouse N-myc gene were constructed, in which codons for the first and second leucine residues within this structure were systematically replaced by other amino acids. The expression plasmids which contained the mutated and wild type N-myc genes were cotransfected into rat embryo cells with activated c-Ha-ras gene and their transforming abilities were compared. It was shown that single amino acid substitutions in the leucine-zipper region inactivate the transforming ability of the N-myc gene product. In particular, proline, which is known to disrupt an alpha-helical structure, completely inactivated the transforming activity even when it was substituted for another amino acid located between these two leucine residues. Among several amino acid species used for substitution of the leucine residues, only methionine was able to retain the transforming activities in both the first and second leucine positions, although the activity was reduced as compared with wild-type N-myc gene product. It also appeared that the integrity of the first leucine is more important than the second leucine. Our results provide experimental evidence for the physiological importance of the hypothetical leucine-zipper structure.
Nakajima
H
H
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Ikeda
M
M
Tsuchida
N
N
Nishimura
S
S
Taya
Y
Y
D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000648:ultrastructure; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics* / Q000648:ultrastructure; D011487:Protein Conformation; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000648:ultrastructure; D016271:Proto-Oncogene Proteins c-myc; D051381:Rats; D013329:Structure-Activity Relationship
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D016271:Proto-Oncogene Proteins c-myc