Localization of a metalloproteinase and its inhibitor in the protein bodies of buckwheat seeds

Abstract

Cotyledons of dry buckwheat (Fagopyrum esculentum Moench) seeds were used to study the cellular localization of a metalloproteinase which performs in vitro the initial limited proteolysis of the main storage protein of the seed, and of its proteinaceous inhibitor. Fractions of complex protein bodies (PB 1) and of the cytoplasm and membrane material (CMM) were obtained by fractionating cotyledons in a mixture of acetone and CCl4. The greater part of the metalloproteinase activity was found to be localized in the PB 1 fraction, with a lesser amount in the CMM fraction, whereas the metalloproteinase inhibitor was localized almost entirely in the PB 1 fraction. The data obtained indicate that the complex protein bodies of dry buckwheat seeds contain the components of the proteolytic system responsible for the initial degradation of the main storage protein - the 13S globulin - of buckwheat seeds, i.e. 13S globulin, the metalloproteinase, and its inhibitor. This confirms that it is possibile for the metalloproteinase to perform a controlled proteolysis of the 13S globulin in vivo. The effect of divalent cations on the degradation of the 13S globulin was also studied. A mechanism is discussed whereby the proteolysis of 13S globulin is initiated by divalent cations released as a result of phytin decationization during seedling growth.

Characterization of the Endoproteases Appearing during Wheat Grain Development

“…The significant inhibition exerted by EDTA shows the presence of metalloproteases in mature wheat grains. Metalloproteases are also detected in mature grains of buckwheat (Belozersky et al, 1990;Elpidina et al, 1991) but not in barley (Wrobel and Jones, 1992). Since in buckwheat seeds metalloproteases are localized in the protein bodies, Elpidina et al (1991) proposed a role for them in the initial limited proteolysis of storage proteins early after imbibition.…”

“…Metalloproteases are also detected in mature grains of buckwheat (Belozersky et al, 1990;Elpidina et al, 1991) but not in barley (Wrobel and Jones, 1992). Since in buckwheat seeds metalloproteases are localized in the protein bodies, Elpidina et al (1991) proposed a role for them in the initial limited proteolysis of storage proteins early after imbibition. Aspartic endoproteases are present in mature seeds of wheat (Morris et al, 1985;Belozersky et al, 1989) and barley (Sarkkinen et al, 1992).…”

Proteolytic activities in Phaseolus vulgaris cotyledons under copper stress

“…Maximum activity (100 %)015.12± 2.03 μg azocasein/min/g FW. The following buffers (0.1 M) were used for indicated pH ranges: Na-citrate, 4 to 5.5; Tris-maleate, 6 to 8.5; glycine, 9 and 9.5 proteases (Nielsen and Liener 1984) as well as MP (Belozersky et al 1990;Elpidina et al 1991) represent the major hydrolytic activities in bean cotyledons and are major factors controlling the degradation of storage proteins in germinating seeds (Shutov and Vaintraub 1987). However, upon Cu treatment, SH-proteases activity dependent abundance decreased dramatically as compared to the control pattern; 21 vs 56 % at 9th day.…”

Ethylene regulates the expression of a cysteine proteinase gene during germination of chickpea (Cicer arietinum L.)

“…According to their active site, proteolytic enzymes are classified as aspartic-acid proteinases, serine proteinases, metallo proteinases and cysteine proteinases [31 ]. Although all four types of proteinases have been purified from seeds [ 11,25,35,36] it is recognized that during germination, there is a marked increase in the activity of cysteine proteinases, which are responsible for the catabolism of the majority of reserve proteins [24,28].…”