Collapse and search dynamics of apomyoglobin folding revealed by submillisecond observations of α-helical content and compactness
- Takanori Uzawa*,
- Shuji Akiyama†,
- Tetsunari Kimura*,
- Satoshi Takahashi*,‡,§,¶,
- Koichiro Ishimori*,
- Isao Morishima*, and
- Tetsuro Fujisawa†,¶
- *Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Nishikyo, Kyoto 615-8510, Japan; †RIKEN Harima Institute/SPring-8, Structural Biochemistry Laboratory, Sayo, Hyogo 679-5148, Japan; and ‡Precursory Research for Embryonic Science and Technology, Japan Science and Technology Corporation, Kawaguchi, Saitama 332-0012, Japan
-
Edited by Robert L. Baldwin, Stanford University Medical Center, Stanford, CA, and approved November 13, 2003 (received for review August 21, 2003)
Abstract
The characterization of protein folding dynamics in terms of secondary and tertiary structures is important in elucidating the features of intraprotein interactions that lead to specific folded structures. Apomyoglobin (apoMb), possessing seven helices termed A–E, G, and H in the native state, has a folding intermediate composed of the A, G, and H helices, whose formation in the submillisecond time domain has not been clearly characterized. In this study, we used a rapid-mixing device combined with circular dichroism and small-angle x-ray scattering to observe the submillisecond folding dynamics of apoMb in terms of helical content (f H) and radius of gyration (R g), respectively. The folding of apoMb from the acid-unfolded state at pH 2.2 was initiated by a pH jump to 6.0. A significant collapse, corresponding to ≈50% of the overall change in R g from the unfolded to native conformation, was observed within 300 μs after the pH jump. The collapsed intermediate has a f H of 33% and a globular shape that involves >80% of all its atoms. Subsequently, a stepwise helix formation was detected, which was interpreted to be associated with a conformational search for the correct tertiary contacts. The characterized folding dynamics of apoMb indicates the importance of the initial collapse event, which is suggested to facilitate the subsequent conformational search and the helix formation leading to the native structure.
Footnotes
-
↵ ¶ To whom correspondence may be addressed. E-mail: st{at}protein.osaka-u.ac.jp or fujisawa{at}spring8.or.jp.
-
↵ § Present address: Institute for Protein Research, Osaka University, Suita, Osaka 565-0871, Japan.
-
This paper was submitted directly (Track II) to the PNAS office.
-
Abbreviations: apoMb, apomyoglobin; SAXS, small-angle x-ray scattering; cyt c, cytochrome c.
- Copyright © 2004, The National Academy of Sciences
