CD measurements were carried out on a spectropolarimeter
(JASCO J-815, Japan). The protein concentrations were: commercial
bromelain (2.5 lM) and purified, cryoprotected bromelain (4.2
lM) in phosphate buffer of pH 7.0 at 25 C. CD spectra were measured
in the far-UV range (190–250 nm) in a 10 mm pathlength
quartz cuvette. The data were averaged for 5 scans performed at
a speed of 50 nm/min, and collected in 0.5 nm steps. The baselines
(buffer alone) were subtracted from the protein spectra. Results
were expressed as mean residue ellipticity, [h], defined as [h] =
hobs/(10 C l n), where hobs is the CD in millidegrees, C is the protein
concentration (M), l is the path-length of the cuvette (cm) and
n is the number of amino acid residues assuming a mean number
of 212 residues.