Mobilization of reserve proteins and activities of cysteine peptidases during germinative and post-germinative events of cowpea seeds.
Cysteine peptidases are the best characterized peptidases among those involved with storage protein mobilization during seed germination. In the present work we show two major groups of cysteine peptidase activities, one of higher (55 to 97 kDa) and other with lower (15 to 20 kDa) molecular masses which are temporally activated after 24 and 48 HAI, respectively, in germinating cowpeas. The former group is found both in protein bodies and in cytoplasmic fraction, while the latter is mostly present outside protein bodies. A third cysteine peptidase activity of ~37 kDa was specifically active at quiescent cotyledons and at 12 and 60 hours after imbibition (HAI). Main peptidase activities of albumin fractions were synchronizedly detected with radicle emergence at 36 HAI. Major vicilin mobilization was more pronounced from 60 HAI onwards and steadily increased until 144 HAI, when low levels of the smallest vicilin subunit were present. Cysteine peptidases were susceptible to iodoacetamide, E-64, iodoacetic acid, pCMB and β-mercaptoethanol, except for the ~37 kDa peptidase, which was not affected by any of the inhibitors. By a two-dimensional native/SDS-PAGE combination it was observed an apparent linear arrangement of protein breakdown products as well as of peptidase activity spots. The finding may indicate a complex set of sequential proteolytic events where peptidases induce or activate new peptidases, which may act upon different aggregates or zymogens, and these hydrolysis products appear in a line of constant decreasing Rf x Mr ratio.
Universidade Estadual do Norte Fluminense Darcy Ribeiro-UENF, Centro de Biociências e Biotecnologia-CBB, Campos dos Goytacazes, RJ, Brazil.
Lucilene Ollivier
LO
de Oliveira
Cláudio Andrés
CA
Retamal
Maura
M
da Cunha
Antônia Elenir Amâncio
AE
Oliveira
Márcio dos Santos Teixeira
Mdos S
Pinto
Kátia Valevski Sales
KV
Fernandes
D000418:Albumins / Q000737:chemistry / Q000378:metabolism; D018548:Cotyledon / Q000737:chemistry* / Q000201:enzymology; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D003593:Cytoplasm / Q000737:chemistry / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D007887:Fabaceae / Q000737:chemistry* / Q000201:enzymology; D018525:Germination; D005916:Globulins / Q000737:chemistry / Q000378:metabolism; D010940:Plant Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*
D000418:Albumins; D005916:Globulins; D010940:Plant Proteins; D003546:Cysteine Endopeptidases