Protein Conformation Changes of HemAT-Bs upon Ligand Binding Probed by Ultraviolet Resonance Raman Spectroscopy

• Published in: The Journal of biological chemistry Vol. 283; no. 11; pp. 6942 - 6949
• Main Authors: El-Mashtoly, Samir F., Gu, Yuzong, Yoshimura, Hideaki, Yoshioka, Shiro, Aono, Shigetoshi, Kitagawa, Teizo
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 14.03.2008; American Society for Biochemistry and Molecular Biology
• Subjects: Bacillus subtilis; Bacillus subtilis - metabolism; Bacterial Proteins - chemistry; Gene Expression Regulation, Bacterial; Heme - chemistry; Heme-Binding Proteins; Hemeproteins - chemistry; Hydrogen Bonding; Ligands; Models, Biological; Models, Molecular; Molecular Conformation; Mutation; Oxygen - metabolism; Protein Conformation; Spectrophotometry, Ultraviolet - methods; Spectrum Analysis, Raman - methods; Tyrosine - chemistry
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M709209200

HemAT from Bacillus subtilis (HemAT-Bs) is a heme-based O2 sensor protein that acts as a signal transducer responsible for aerotaxis. HemAT-Bs discriminates its physiological effector (O2) from other gas molecules (CO and NO), although all of them bind to a heme. To monitor the conformational changes in the protein moiety upon binding of different ligands, we have investigated ultraviolet resonance Raman (UVRR) spectra of the ligand-free and O2-, CO-, and NO-bound forms of full-length HemAT-Bs and several mutants (Y70F, H86A, T95A, and Y133F) and found that Tyr70 in the heme distal side and Tyr133 and Trp132 from the G-helix in the heme proximal side undergo environmental changes upon ligand binding. In addition, the UVRR results confirmed our previous model, which suggested that Thr95 forms a hydrogen bond with heme-bound O2, but Tyr70 does not. It is deduced from this study that hydrogen bonds between Thr95 and heme-bound O2 and between His86 and heme 6-propionate communicate the heme structural changes to the protein moiety upon O2 binding but not upon CO and NO binding. Accordingly, the present UVRR results suggest that O2 binding to heme causes displacement of the G-helix, which would be important for transduction of the conformational changes from the sensor domain to the signaling domain.