Truncation of a cross-linked GCN4-p1 coiled coil leads to ultrafast folding.
Structural perturbation has been extensively used in protein folding studies because it yields valuable conformational information regarding the folding process. Here we have used N-terminal truncation on a cross-linked variant of the GCN4-p1 leucine zipper, aiming to develop a better understanding of the folding mechanism of the coiled-coil motif. Our results indicate that removing the first heptad repeat in this cross-linked GCN4-p1 coiled coil significantly decreases the folding free energy barrier and results in a maximum folding rate of (2.0 +/- 0.3 micros)(-1), which is approximately 50 times faster than that of the full-length protein. Therefore, these results suggest that a set of native or nativelike tertiary interactions, distributed throughout the entire sequence, collectively stabilize the folding transition state of the GCN4-p1 coiled coil. While stable subdomains or triggering sequences have been shown to be critical to the stability of GCN4 coiled coils, our results suggest that the folding of such a subdomain does not seem to dictate the overall folding kinetics.
Bunagan
Michelle R
MR
Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Cristian
Lidia
L
DeGrado
William F
WF
Gai
Feng
F
D000595:Amino Acid Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D002942:Circular Dichroism; D003432:Cross-Linking Reagents / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D016350:Leucine Zippers; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013055:Spectrophotometry, Infrared; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
D050976:Basic-Leucine Zipper Transcription Factors; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; C501391:GCN4 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors