title: Using nitrile-derivatized amino acids as infrared probes of local environment.
abstract: It is well-known that the C=N stretching vibration in acetonitrile is sensitive to solvent. Therefore, we proposed in this contribution to use this vibrational mode to report local environment of a particular amino acid in proteins or local environmental changes upon binding or folding. We have studied the solvent-induced frequency shift of two nitrile-derivatized amino acids, which are, AlaCN and PheCN, in H(2)O and tetrahydrofuran (THF), respectively. Here, THF was used to approximate a protein's hydrophobic interior because of its low dielectric constant. As expected, the C=N stretching vibrations of both AlaCN and PheCN shift as much as approximately 10 cm(-1) toward higher frequency when THF was replaced with H2O, indicative of the sensitivity of this vibration to solvation. To further test the utility of nitrile-derivatized amino acids as probes of the environment within a peptide, we have studied the binding between calmodulin (CaM) and a peptide from the CaM binding domain of skeletal muscle myosin light chain kinase (MLCK(579-595)), which contains a single PheCN. MLCK(579-595) binds to CaM in a helical conformation. When the PheCN was substituted on the polar side of the helix, which was partially exposed to water, the C=N stretching vibration is similar to that of PheCN in water. In constrast, when PheCN is introduced at a site that becomes buried in the interior of the protein, the C=N stretch is similar to that of PheCN in THF. Together, these results suggest that the C=N stretching vibration of nitrile-derivatized amino acids can indeed be used as local internal environmental markers, especially for protein conformational studies.
authors.affiliation: Department of Chemistry, University of Pennsylvania, Philadelphia 19104, USA.
authors.forename: Zelleka
authors.initials: Z
authors.lastname: Getahun
authors.forename: Cheng-Yen
authors.initials: CY
authors.lastname: Huang
authors.forename: Ting
authors.initials: T
authors.lastname: Wang
authors.forename: Brenda
authors.initials: B
authors.lastname: De León
authors.forename: William F
authors.initials: WF
authors.lastname: DeGrado
authors.forename: Feng
authors.initials: F
authors.lastname: Gai
mesh_terms: D000409:Alanine / Q000031:analogs & derivatives*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D002147:Calmodulin / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009219:Myosin-Light-Chain Kinase / Q000737:chemistry / Q000378:metabolism; D009570:Nitriles / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010649:Phenylalanine / Q000031:analogs & derivatives*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*
chemical_list: D000596:Amino Acids; D002147:Calmodulin; D009570:Nitriles; D010446:Peptide Fragments; D010649:Phenylalanine; D009219:Myosin-Light-Chain Kinase; D000409:Alanine