PLANT PHYSIOLOGY , Vol 104, Issue 2 401-407, Copyright © 1994 by American Society of Plant Biologists
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DEVELOPMENT AND GROWTH REGULATION |
Development of Endopeptidase Activities in Maize (Zea mays L.) Endosperms
W. Mitsuhashi and A. Oaks
Department of Botany, University of Guelph, Guelph, Ontario, Canada N1G 2W1
An activity stain was used after native polyacrylamide gel electrophoresis,
and at least 17 different endopeptidase activities were detected in maize
(Zea mays L.) endosperm extracts prepared during the first 6 d after
imbibition. The enzymes detected were classified into four groups based on
their time of appearance and on their mobility in polyacrylamide gels. The
first group, which included two enzymes present in dry endosperms,
disappeared soon after imbibition. The second group, comprising five
activity bands, appeared during the first 2 to 3 d after imbibition and
then disappeared. The third set of enzymes increased continuously
throughout the experimental period. The fourth group appeared after d 3 and
remained at a constant level after that time. The endopeptidase activities
were characterized by the effect of specific inhibitors on their
activities. The two enzymes of the first group are metalloendopeptidases
based on their sensitivity to ethylenediaminetetracetate (EDTA). Enzymes of
the second, third, and fourth groups are sulfhydryl-endopeptidases as
judged by their sensitivity to antipain, chymostatin, leupeptin, and E-64
and by their requirement for 2-mercaptoethanol. Pepstatin,
phenylmethylsulfonyl fluoride, or EDTA had no effect on these enzymes. Many
of the second, third, and fourth group enzymes cleaved [alpha]-zein-rich
proteins as well as such easily obtained proteins as gelatin (used in our
standard assay) and hemoglobin. The second group had a high affinity for
[gamma]-zein, whereas none of the bands in the fourth group of enzymes
cleaved this type of zein. The two metalloenzymes of the first group
cleaved neither [alpha]- nor [gamma]-zeins.
