title: Steady-state and time-resolved fluorescence spectroscopic studies on interaction of the N-terminal region with the hairpin loop of the phytocystatin Scb.
abstract: The steady-state and time-resolved fluorescence spectroscopy is one of the most powerful method to detect and analyze subtle conformation change and interaction between peptide elements in protein. Phytocystatin Scb isolated from sunflower seeds includes a single Trp residue at position 85. In an attempt to investigate the interaction of the N-terminal region of Scb with the first and second hairpin loops by fluorescence spectroscopy of Trp residue, two Scb mutants in which single Trp locates at position 52 and 58, respectively, and their N-terminal removed mutants were generated. The N-terminal truncation changed the fluorescence decay kinetics of Trp52 from the triple exponential to double. Furthermore, the time-resolved fluorescence anisotropy residue indicated that the segmental motion of Trp52 was significantly enhanced by its N-terminal truncation. In contrast, Trp58 and Trp85 had little influence. The N-terminal successive truncations of Scb and its mutants resulted in the weaken inhibitors to papain. These results suggested that the N-terminal region of Scb interacts with the peptide segment preceding the first hairpin loop, thereby stabilizing the conformation of the hairpin loop structure.
authors.affiliation: Laboratory of Biochemistry, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School of Kyushu University, Hakozaki, Fukuoka 812-8581, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Doi-Kawano
authors.forename: Etsuko
authors.initials: E
authors.lastname: Nishimoto
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Kouzuma
authors.forename: Daisuke
authors.initials: D
authors.lastname: Takahashi
authors.forename: Shoji
authors.initials: S
authors.lastname: Yamashita
authors.forename: Makoto
authors.initials: M
authors.lastname: Kimura
mesh_terms: D015891:Cystatins / Q000737:chemistry* / Q000302:isolation & purification; D006368:Helianthus / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012639:Seeds / Q000737:chemistry; D013050:Spectrometry, Fluorescence; D013997:Time Factors
chemical_list: D015891:Cystatins; D010455:Peptides