An investigation into the native-like properties of de novo designed cavitand-based four-helix bundle proteins.
We investigated the hydrophobic packing of two previously designed caviteins, LG2 and LG3, which differ by one Gly in the linker regions between the peptide sequence and the cavitand scaffold. We sought to diminish the putative native-like properties of LG2 and LG3, and see if we could diagnose a change in the conformational specificity of the hydrophobic core. We replaced the leucine residues with norleucine residues at the hydrophobic positions in LG2 and LG3, to create NG2 and NG3, respectively. LG2 exhibited more dispersion, but less sharp signals than LG3 in the amide region of its (1)H NMR spectrum. NG3 and NG2 were found to be slightly less helical and significantly less stable toward guanidine hydrochloride compared with their reference caviteins. The (1)H NMR spectrum of NG2 was very similar to that of LG2, whereas there was a noticeable loss in the number and sharpness of the amide signals of NG3 compared with LG3. These data suggest that LG3 is very well packed; a loss in conformational specificity resulted from replacement of the leucine residues with norleucine residues. In contrast, the packing and dynamics of the hydrophobic core in LG2 were similar to those in NG2 (both more modest than LG3), as their (1)H NMR spectra were virtually indistinguishable. Overall, substitution of leucine by norleucine provided an efficient, convenient, and informative probe of the packing and dynamics of our caviteins' hydrophobic cores.
Huttunen-Hennelly
Heidi E K
HE
Department of Chemistry, Thompson Rivers University, 900 McGill Road, Kamloops, BC, Canada.
Sherman
John C
JC
D002942:Circular Dichroism; D004988:Ethers, Cyclic / Q000138:chemical synthesis / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D002727:Proguanil / Q000737:chemistry; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D012118:Resorcinols / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet
D004988:Ethers, Cyclic; D010455:Peptides; D012118:Resorcinols; C442939:cavitand; C527838:cavitein LG2; C527839:cavitein LG3; D002727:Proguanil