title: Role of protein surface charge in monellin sweetness.
abstract: A small number of proteins have the unusual property of tasting intensely sweet. Despite many studies aimed at identifying their sweet taste determinants, the molecular basis of protein sweetness is not fully understood. Recent mutational studies of monellin have implicated positively charged residues in sweetness. In the present work, the effect of overall net charge was investigated using the complementary approach of negative charge alterations. Multiple substitutions of Asp/Asn and Glu/Gln residues radically altered the surface charge of single-chain monellin by removing six negative charges or adding four negative charges. Biophysical characterization using circular dichroism, fluorescence, and two-dimensional NMR demonstrates that the native fold of monellin is preserved in the variant proteins under physiological solution conditions although their stability toward chemical denaturation is altered. A human taste test was employed to determine the sweetness detection threshold of the variants. Removal of negative charges preserves monellin sweetness, whereas added negative charge has a large negative impact on sweetness. Meta-analysis of published charge variants of monellin and other sweet proteins reveals a general trend toward increasing sweetness with increasing positive net charge. Structural mapping of monellin variants identifies a hydrophobic surface predicted to face the receptor where introduced positive or negative charge reduces sweetness, and a polar surface where charges modulate long-range electrostatic complementarity.
authors.affiliation: Department of Biophysical Chemistry, Lund University, Lund, Sweden.
authors.forename: Wei-Feng
authors.initials: WF
authors.lastname: Xue
authors.forename: Olga
authors.initials: O
authors.lastname: Szczepankiewicz
authors.forename: Eva
authors.initials: E
authors.lastname: Thulin
authors.forename: Sara
authors.initials: S
authors.lastname: Linse
authors.forename: Jannette
authors.initials: J
authors.lastname: Carey
mesh_terms: D000328:Adult; D000368:Aged; D019943:Amino Acid Substitution; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D043562:Receptors, G-Protein-Coupled / Q000502:physiology; D013549:Sweetening Agents / Q000737:chemistry*; D013649:Taste / Q000187:drug effects
chemical_list: D010940:Plant Proteins; D043562:Receptors, G-Protein-Coupled; D013549:Sweetening Agents; C002541:monellin protein, Dioscoreophyllum cumminsii