Co-operative ionisation of aspartic-acid-158 and histidine-159 in papain. Evidence from 19F nuclear-magnetic-resonance and fluorescence spectroscopy.
The chemical shift of the single resonance in the 19F nuclear magnetic resonance spectrum of papain which has been irreversibly inhibited by 3-bromo-1,1,1-trifluoropropanone, exhibits pH-dependence. The fluorescence intensity of this papain derivative shows pH-dependence on two groups which exhibit co-operative ionisation. This co-operative behaviour is probably a function of the probe since the fluorescence intensity of S-ethane-thio-papain is dependent on a single ionisation constant, whereas that of S-(2-hydroxyethane)-thio-papain is dependent on two ionisable groups again acting co-operatively. The 1,1,1-trifluoroketone probe will be hydrated in aqueous solution and would be capable of hydrogen bonding with the protein. The two groups detected are considered to be aspartic-acid-158 and histidine-159. The co-operative ionisation of these groups in substrate hydrolysis is discussed.
D001224:Aspartic Acid; D055598:Chemical Phenomena; D002621:Chemistry; D005461:Fluorine; D006639:Histidine; D006868:Hydrolysis; D066298:In Vitro Techniques; D007477:Ions; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D010206:Papain; D013050:Spectrometry, Fluorescence; D013438:Sulfhydryl Compounds / Q000138:chemical synthesis
D007477:Ions; D013438:Sulfhydryl Compounds; D005461:Fluorine; D001224:Aspartic Acid; D006639:Histidine; D010206:Papain