title: Structural role of the terminal disulfide bond in the sweetness of brazzein.
abstract: Brazzein, a 54 residue sweet-tasting protein, is thought to participate in a multipoint binding interaction with the sweet taste receptor. Proposed sites for interaction with the receptor include 2 surface loops and the disulfide bond that connects the N- and C-termini. However, the importance of each site is not well understood. To characterize the structural role of the termini in the sweetness of brazzein, the position of the disulfide bond connecting the N- and C-termini was shifted by substituting K3-C4-K5 with C3-K4-R5. The apparent affinity and V(max) of the C3-K4-R5-brazzein (CKR-brazzein) variant were only modestly decreased compared with the wild-type (WT) brazzein. We determined a high-resolution structure of CKR-brazzein by nuclear magnetic resonance spectroscopy (backbone root mean square deviation of 0.39 Å). Comparing the structure of CKR-brazzein with that of WT-brazzein revealed that the terminal β-strands of the variant display extended β-structure and increased dynamics relative to WT-brazzein. These results support previous mutagenesis studies and further suggest that, whereas interactions involving the termini are necessary for full function of brazzein, the termini do not constitute the primary site of interaction between brazzein and the sweet taste receptor.
authors.affiliation: Biochemistry Department, University of Wisconsin-Madison, USA.
authors.forename: Sannali M
authors.initials: SM
authors.lastname: Dittli
authors.forename: Hongyu
authors.initials: H
authors.lastname: Rao
authors.forename: Marco
authors.initials: M
authors.lastname: Tonelli
authors.forename: Jeniffer
authors.initials: J
authors.lastname: Quijada
authors.forename: John L
authors.initials: JL
authors.lastname: Markley
authors.forename: Marianna
authors.initials: M
authors.lastname: Max
authors.forename: Fariba
authors.initials: F
authors.lastname: Assadi-Porter
authors.forename: Emeline
authors.initials: E
authors.lastname: Maillet
mesh_terms: D004220:Disulfides / Q000737:chemistry*; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D015394:Molecular Structure; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013549:Sweetening Agents / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013649:Taste
chemical_list: D004220:Disulfides; D010940:Plant Proteins; D013549:Sweetening Agents; C090521:brazzein protein, Pentadiplandra brazzeana