title: Folding nuclei of the scFv fragment of an antibody.
abstract: The folding kinetics of the variable domains of the phosphorylcholine-binding antibody McPC603, combined into a scFv fragment [VH-(Gly4Ser)3-VL], were investigated by the use of fluorescence spectroscopy, nuclear magnetic resonance (NMR), and mass spectrometry (MS). All three methods gave evidence for the occurrence of a major kinetic intermediate during the refolding of the denatured, oxidized scFv fragment. This intermediate is formed within the first 30 s of folding and comprises exchange-protected amide protons of hydrophobic and aromatic amino acids, most of which are localized within the inner beta-sheet of the V(L) domain. In the subsequent slow step, most of the amide protons become protected with rate constants that are very similar for residues of both domains. These data are in agreement with the MS results, which indicate a cooperative folding event from the intermediate to the native state of the scFv fragment.
authors.affiliation: Biochemisches Institut, Universität Zürich, Switzerland.
authors.forename: C
authors.initials: C
authors.lastname: Freund
authors.forename: A
authors.initials: A
authors.lastname: Honegger
authors.forename: P
authors.initials: P
authors.lastname: Hunziker
authors.forename: T A
authors.initials: TA
authors.lastname: Holak
authors.forename: A
authors.initials: A
authors.lastname: Plückthun
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007135:Immunoglobulin Variable Region / Q000737:chemistry*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D013050:Spectrometry, Fluorescence
chemical_list: D000906:Antibodies; D007140:Immunoglobulin Fab Fragments; D007135:Immunoglobulin Variable Region; C069242:McPC603 antibody