The mutability of stem bromelain: evidence for perturbation by structural transitions of the parameters that characterize the reaction of the essential thiol group of bromelain with 2,2′-dipyridyl disulphide

Stem bromelain (bromelain, EC 3.4.4.24) re- sembles papain and ficin in some respects (see Murachi, 1970; Murachi & Takahashi, 1970), in particular in that it contains an essential cysteine residue, the thiol group of which is within 0.5nm (5A) of the imidazole group of a histidine residue (Husain & Lowe, 1968). Bromelain differs from the other two enzymes, however, notably in some of its catalytic properties (see Brocklehurst et al., 1967) and in that it is a glycoprotein (Murachi et al., 1967). Previous work in this laboratory (Wharton, 1969) had demonstrated a marked difference in the catalytic characteristics (see below) of crude and purified bromelain preparations. Our finding that 2,2'- dipyridyl disulphide may be used to detect nucleo- philic reactivity in the imidazole-thiol pair of papain (Brocklehurst & Little, 1972) led us to investigate the reaction of this reagent with bromelain. The objects were the detection of (i) a similar nucleophilic system in bromelain and (ii), if this exists, any change in reactivity that the system might undergo during the enzyme purification. Most of the recorded properties of bromelain relate to bromelain prepared essentially by the method of Murachi (Murachi et al., 1964; Murachi, 1970). This method includes chromato- graphy on a cation-exchange resin such as Amberlite CG-50. The product is less than fully active, and usually an 'activator' such as L-cysteine, KCN or dithiothreitol is added to the purified enzyme.