title: A spectroscopic and conformational study of pertussis toxin.
abstract: The conformation of native pertussis toxin has been investigated by secondary structure prediction and by circular dichroism, fluorescence and second-derivative ultraviolet absorption spectroscopy. The far-ultraviolet circular dichroic spectrum is characteristic of a protein of high beta-sheet and low alpha-helix content. This is also shown by an analysis of the circular dichroic spectrum with the Contin programme which indicates that the toxin possesses 53% beta-sheet, 10% alpha-helix and 37% beta-turn/loop secondary structure. Second-derivative ultraviolet absorption spectroscopy suggests that 34 tyrosine residues are solvent-exposed and quenching of tryptophan fluorescence emission has shown that 4 tryptophan residues are accessible to iodide ions. One of these tryptophans appears to be in close proximity to a positively charged side-chain, since only 3 tryptophans are accessible to caesium ion fluorescence quenching. When excited at 280 nm, the emission spectrum contains a significant contribution from tyrosine fluorescence, which may be a consequence of the high proportion (55%) of surface-exposed tyrosines. No changes in the circular dichroic spectra of the toxin were found in the presence of the substrate NAD. However, NAD did quench both tyrosine and tryptophan fluorescence emission but did not change the shape of the emission spectrum, or the accessibility of the tryptophans to either the ionic fluorescence quenchers or the neutral quencher acrylamide.
authors.affiliation: Division of Biotechnology, Public Health Laboratory Service, Porton Down, Salisbury, England.
authors.forename: R N
authors.initials: RN
authors.lastname: Seabrook
authors.forename: T
authors.initials: T
authors.lastname: Atkinson
authors.forename: L I
authors.initials: LI
authors.lastname: Irons
mesh_terms: D002942:Circular Dichroism; D007700:Kinetics; D009243:NAD / Q000494:pharmacology; D037342:Pertussis Toxin; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D010566:Virulence Factors, Bordetella / Q000737:chemistry*
chemical_list: D010566:Virulence Factors, Bordetella; D009243:NAD; D037342:Pertussis Toxin