Structural study on acid-induced unfolding intermediates of myoglobin by using UV resonance Raman scattering from tryptophan residues

✍ Scribed by Shinji Hashimoto; Jyun-ichi Fukasaka; Hideo Takeuchi

• Book ID: 102430615
• Publisher: John Wiley and Sons
• Year: 2001
• Tongue: English
• Weight: 158 KB
• Volume: 32
• Category: Article
• ISSN: 0377-0486
• DOI: 10.1002/jrs.729

✦ Synopsis

Abstract

Ultraviolet resonance Raman spectra of sperm whale myoglobin were examined at various pH in the presence and absence of a helix‐stabilizing anion, SO~4~^2−^. Sperm whale myoglobin contains two Trp residues (Trp7 and Trp14) in helix A. The 229 nm excited Raman spectra of metmyoglobin and apomyoglobin in the native form at pH 7 show that these Trp residues are located in hydrophobic environments and their indole nitrogen atoms are hydrogen bonded to proton acceptors, presumably to water molecules. The Raman intensity of Trp decreases in two successive steps as the pH is lowered. The intensity decrease in the transition from the native form to an unfolding intermediate at pH 4 is ascribable to a decrease in environmental hydrophobicity of Trp14 caused by unfolding of helix E, from which the hydrophobic side‐chains of three Leu residues protrude toward Trp14 in the native structure. Addition of 30 mM SO~4~^2−^ to a solution of the unfolding intermediate results in a marked change of the Trp14 side‐chain torsion angle, |χ^2,1^|, as evidenced by a wavenumber shift and an increase in bandwidth of the W3 band of Trp. The fully unfolded form at pH 2 is partially refolded by the binding of SO~4~^2−^. Although the secondary structure of the refolded form is similar to that of the unfolding intermediate, the tertiary structure differs between the two forms. Ultraviolet resonance Raman scattering from Trp residues is a good probe for protein structure in the folding or unfolding pathway. Copyright © 2001 John Wiley & Sons, Ltd.