title: Synthetic interface peptides as inactivators of multimeric enzymes: inhibitory and conformational properties of three fragments from Lactobacillus casei thymidylate synthase.
abstract: Three synthetic peptides corresponding to distinct segments of the subunit interface of the dimeric enzyme thymidylate synthase (residues 17-38, N 22; residues 174-190, M 17; and residues 201-220, C 20) have been investigated for their ability to function as inhibitors by modifying the quaternary structure of the enzyme. A dramatic reduction of enzyme activity is observed following incubation of TS with the C 20 peptide. The N 22 and M 17 peptides were unable to cause any loss of enzymatic activity. Addition of the C 20 peptide results in a loss of fluorescence of TS labeled with a dansyl group at Cys 198, following aggregation and precipitation of the protein. The effects are not observed for the N 22 or M 17 peptides. Loss of enzymatic activity is related to the ability of C 20 to promote protein aggregation. The conformations of the peptides have been studied using CD and NMR in order to correlate the observed function with solution structures. Peptides N 22 and M 17 are largely unstructured in aqueous solution. A population of nascent helical structures or multiple turn conformations has been detected for the C 20 peptide in aqueous solution by NMR. Addition of 50% (v/v) hexafluoroacetone trihydrate (HFA), a structure-stabilizing cosolvent, stabilizes the helical conformation in the C 20 peptide. Under similar conditions, N 22 and M 17 remain largely extended with observations of local beta-turn conformations. Interestingly, the C 20 peptide is a beta-hairpin in the native structure, whereas the other two peptides are individual strand components of a beta-sheet.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore.
authors.forename: V
authors.initials: V
authors.lastname: Prasanna
authors.forename: S
authors.initials: S
authors.lastname: Bhattacharjya
authors.forename: P
authors.initials: P
authors.lastname: Balaram
mesh_terms: D000096:Acetone / Q000031:analogs & derivatives; D000595:Amino Acid Sequence; D002384:Catalysis; D002942:Circular Dichroism; D019281:Dimerization; D004791:Enzyme Inhibitors / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D005466:Fluorocarbons; D007780:Lacticaseibacillus casei / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology*; D011487:Protein Conformation / Q000187:drug effects; D012996:Solutions; D013940:Thymidylate Synthase / Q000737:chemistry* / Q000494:pharmacology*; D014867:Water; D006845:Hydrocarbons, Fluorinated
chemical_list: D000096:Acetone; D004791:Enzyme Inhibitors; D005466:Fluorocarbons; D010446:Peptide Fragments; D012996:Solutions; D013940:Thymidylate Synthase; D014867:Water; C017366:hexafluoroacetone; D006845:Hydrocarbons, Fluorinated