title: Fluorous effect in proteins: de novo design and characterization of a four-alpha-helix bundle protein containing hexafluoroleucine.
abstract: Several studies have demonstrated that proteins incorporating fluorinated analogues of hydrophobic amino acids such as leucine and valine into their hydrophobic cores exhibit increased stability toward thermal denaturation and unfolding by guanidinium chloride. However, estimates for the increase in the thermodynamic stability of a protein (DeltaDeltaG(unfold)) afforded by the substitution of a hydrophobic amino acid with its fluorinated analogue vary quite significantly. To address this, we have designed a peptide that adopts an antiparallel four-helix bundle structure in which the hydrophobic core is packed with leucine, and investigated the effects of substituting the central two layers of the core with L-5,5,5,5',5',5'-hexafluoroleucine (hFLeu). We find that DeltaDeltaG(unfold) is increased by 0.3 kcal/mol per hFLeu residue. This is in good agreement with the predicted increase in DeltaDeltaG(unfold) of 0.4 kcal/mol per residue arising from the increased hydrophobicity of the hFLeu side chain, which we determined experimentally from partitioning measurements on hFLeu and leucine. The increased stability of this fluorinated protein may therefore be ascribed to simple hydrophobic effects, rather than specific "fluorous" interactions between the hFLeu residues.
authors.affiliation: Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109-1055, USA.
authors.forename: Kyung-Hoon
authors.initials: KH
authors.lastname: Lee
authors.forename: Hyang-Yeol
authors.initials: HY
authors.lastname: Lee
authors.forename: Morris M
authors.initials: MM
authors.lastname: Slutsky
authors.forename: James T
authors.initials: JT
authors.lastname: Anderson
authors.forename: E Neil G
authors.initials: EN
authors.lastname: Marsh
mesh_terms: D002942:Circular Dichroism; D019791:Guanidine / Q000378:metabolism; D007930:Leucine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D011489:Protein Denaturation; D015202:Protein Engineering; D017433:Protein Structure, Secondary; D013194:Staining and Labeling
chemical_list: C445784:hexafluoroleucine; D007930:Leucine; D019791:Guanidine