Stabilization of coiled-coil peptide domains by introduction of trifluoroleucine.
Substitution of leucine residues by 5,5,5-trifluoroleucine at the d-positions of the leucine zipper peptide GCN4-p1d increases the thermal stability of the coiled-coil structure. The midpoint thermal unfolding temperature of the fluorinated peptide is elevated by 13 degrees C at 30 microM peptide concentration. The modified peptide is more resistant to chaotropic denaturants, and the free energy of folding of the fluorinated peptide is 0.5-1.2 kcal/mol larger than that of the hydrogenated form. A similarly fluorinated form of the DNA-binding peptide GCN4-bZip binds to target DNA sequences with affinity and specificity identical to those of the hydrogenated form, while demonstrating enhanced thermal stability. Molecular dynamics simulation on the fluorinated GCN4-p1d peptide using the Surface Generalized Born implicit solvation model revealed that the coiled-coil binding energy is 55% more favorable upon fluorination. These results suggest that fluorination of hydrophobic substructures in peptides and proteins may provide new means of increasing protein stability, enhancing protein assembly, and strengthening receptor-ligand interactions.
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.
Y
Y
Tang
G
G
Ghirlanda
N
N
Vaidehi
J
J
Kua
D T
DT
Mainz
W A
WA
Goddard III
W F
WF
DeGrado
D A
DA
Tirrell
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism*; D007930:Leucine / Q000031:analogs & derivatives* / Q000737:chemistry*; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins; D055672:Static Electricity; D013816:Thermodynamics
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D010455:Peptides; D029701:Saccharomyces cerevisiae Proteins; C035177:5',5',5'-trifluoroleucine; D004247:DNA; D011494:Protein Kinases; D007930:Leucine