The kinetics of the acid-induced conformational change of influenza virus haemagglutinin can be followed using 1,1'-bis(4-anilino-5-naphthalenesulphonic acid).
1,1'-Bis(4-anilino-5-naphthalenesulphonic acid) (bis-ANS) has been shown by fluorescence spectroscopy to bind to bromelain-cleaved influenza haemagglutinin (BHA). The fluorescence intensity of 1.2 microM bis-ANS in the presence of BHA in its low-pH conformation is twenty-fold higher than in the presence of BHA in its neutral-pH conformation. The use of this probe provides a sensitive method for investigating the kinetics of the irreversible conformational change of BHA induced by low pH. At pH5.0 the reaction is described by a rapid burst followed by a double exponential increase in the fluorescence of bis-ANS, with rate constants of 5.2 +/- 0.9 x 10(-3) sec-1 and 6.7 +/- 1.9 x 10(-4) sec-1. This reaction is sensitive to the presence of tert-butylhydroquinone, an inhibitor of the conformational transition of BHA. The dependence of the reaction rate on pH indicates that the acid-induced conformational change is dependent upon the multiple protonation of the neutral-pH conformation of BHA.
Bethell
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RC
Department of Virology, Glaxo Research and Development Ltd., Greenford, Middlesex, U.K.
Gray
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NM
Penn
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CR
D000817:Anilino Naphthalenesulfonates; D000975:Antioxidants / Q000494:pharmacology*; D001963:Bromelains; D005456:Fluorescent Dyes; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D006873:Hydroquinones / Q000494:pharmacology; D007700:Kinetics; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014759:Viral Envelope Proteins / Q000737:chemistry
D000817:Anilino Naphthalenesulfonates; D000975:Antioxidants; D005456:Fluorescent Dyes; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D006389:Hemagglutinins, Viral; D006873:Hydroquinones; D010446:Peptide Fragments; D014759:Viral Envelope Proteins; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); D001963:Bromelains; C018855:2-tert-butylhydroquinone