Structure-based design of a leucine zipper protein with new DNA contacting region.
We have employed a structure-based design to construct a small folding domain from the F-actin bundling protein villin that contains the amino acids necessary for the DNA binding of the basic leucine zipper protein GCN4 and have compared its DNA binding with GCN4. The monomeric motif folds into a stable domain and binds DNA in a rigid-body mechanism, while its affinity is not higher than that of the basic region peptide. The addition of the leucine zipper region to the folded domain restored its sequence-specific DNA binding comparable to that of GCN4. Unlike the monomeric folded domain, its leucine zipper derivative undergoes a conformational change upon DNA binding. CD spectral and thermodynamic studies indicate that the DNA-contacting region is folded in the presence or absence of DNA and suggest that the junction between the DNA-contacting and the leucine zipper regions transits to a helix in the presence of DNA. These results demonstrate that the structural transition outside the direct-contacting region, which adjusts the precise location of the DNA-contacting region, plays a critical role in the specific complex formation of basic leucine zipper proteins.
Morii
Takashi
T
Institute of Advanced Energy, Kyoto University, and PRESTO, Japan Science and Technology Corporation, Uji, Kyoto 611-0011, Japan. t-motii@iae.kyoto-u.ac.jp
Sato
Shin-ichi
S
Hagihara
Masaki
M
Mori
Yasuo
Y
Imoto
Keiji
K
Makino
Keisuke
K
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000138:chemical synthesis / Q000737:chemistry; D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry*; D004268:DNA-Binding Proteins / Q000138:chemical synthesis* / Q000737:chemistry; D019281:Dimerization; D005656:Fungal Proteins / Q000138:chemical synthesis / Q000737:chemistry; D016350:Leucine Zippers; D008840:Microfilament Proteins / Q000138:chemical synthesis / Q000737:chemistry; D008969:Molecular Sequence Data; D010452:Peptide Biosynthesis; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D011494:Protein Kinases / Q000138:chemical synthesis / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D008840:Microfilament Proteins; D029701:Saccharomyces cerevisiae Proteins; C026615:villin; D004247:DNA; D011494:Protein Kinases