Conformational changes during apoplastocyanin folding observed by photocleavable modification and transient grating

Shun Hirota, Yukari Fujimoto, Jungkwon Choi, Naoki Baden, Noriko Katagiri, Masako Akiyama, Rinske Hulsker, Marcellus Ubbink, Toshihide Okajima, Teruhiro Takabe, Noriaki Funasaki, Yoshihito Watanabe, Masahide Terazima

Research output: Contribution to journalArticlepeer-review

26 Citations (Scopus)


A new method to investigate the initial protein folding dynamics is developed based on a pulsed laser light triggering method and a unique transient grating method. The side chain of the cysteine residue of apoplastocyanin (apoPC) was site-specifically modified with a 4,5-dimethoxy-2-nitrobenzyl derivative, where the CD and 2D NMR spectra showed that the modified apoPC was unfolded. The substituent was cleaved with a rate of about 400 ns by photoirradiation, which was monitored by the disappearance of the absorption band at 355 nm and the increase in the transient grating signal. After a sufficient time from the photocleavage reaction, the CD and NMR spectra showed that the native β-sheet structure was recovered. Protein folding dynamics was monitored in the time domain with the transient grating method from a viewpoint of the molecular volume change and the diffusion coefficient, both of which reflect the global structural change, including the protein-water interaction. The observed volume decrease of apoPC with a time scale of 270 μs is ascribed to the initial hydrophobic collapse. The increase in the diffusion coefficient (23 ms) is considered to indicate a change from an intermolecular to an intramolecular hydrogen bonding network. The initial folding process of apoPC is discussed based on these observations.

Original languageEnglish
Pages (from-to)7551-7558
Number of pages8
JournalJournal of the American Chemical Society
Issue number23
Publication statusPublished - 2006 Jun 14
Externally publishedYes

ASJC Scopus subject areas

  • Catalysis
  • General Chemistry
  • Biochemistry
  • Colloid and Surface Chemistry


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