A new serine protease from the latex of
Ipomoea carnea spp
. fistulosa (Morning glory), belonging tothe Convolvulaceae family, was purified to homogeneity by ammonium sulfate fractionation followedby cation exchange chromatography. The enzyme, named carnein, has a molecular mass of 80.24
kDa (matrix-assisted laser desorption/ionization time-of-flight) and an isoelectric point of pH 5.6. ThepH and temperature optima for proteolytic activity were 6.5 and 65
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C, respectively. The extinctioncoefficient (
2801%) of the enzyme was estimated as 37.12, and the protein molecule consists of
35tryptophan, 76 tyrosine, and seven cysteine residues. The effect of several inhibitors such as iodoaceticacid, diisopropylfluorophosphate, phenyl-methanesulfonyl fluoride, chymostatin, soybean trypsininhibitor, HgCl
2, 3
S-3-(
N-{(
S)-1-[
N-(4-guanidinobutyl)carbamoyl]3-ethylbutyl}carbamoyl)oxirane-2-carboxylic acid,
N-ethyl maleimide, ethylene glycol-bis(
![](/images/gifchars/alpha.gif)
-amino ethyl ether)tetraacetic acid, ethylenediamminetetraacetic acid, and
o-phenonthroline indicates that carnein belongs to the family ofserine proteases. The enzyme is not prone to autolysis even at very low concentrations. The N-terminalsequence of carnein (T-T-H-S-P-E-F-L-G-L-A-E-S-S-G-L-X-P-N-S) exhibited considerable similarityto those of other plant serine proteases; the highest similarity was with alnus AG12, one of the subtilasefamily endopepetidases.