Characterization of a soybean beta-conglycinin-degrading protease cleavage site.
Abstract
Protease C1, an enzyme from soybean (Glycine max [L.] Merrill cv Amsoy 71) seedling cotyledons, was previously determined to be the enzyme responsible for the initial degradation of the alpha' and alpha subunits, but not the beta subunit, of beta-conglycinin storage protein. The sizes of the proteolytic products generated by the action of protease C1 suggest that the cleavage sites on the alpha' and alpha subunits of beta-conglycinin may be located in their N-terminal domain, which is not found in the beta subunit of beta-conglycinin. To check this hypothesis, storage proteins from other plant species that are homologous to either the alpha'/alpha or the beta subunit of beta-conglycinin were tested as substrates. As expected, the convicilin from pea (Pisum sativum), a protein homologous to the alpha' and alpha subunits of beta-conglycinin, was digested by protease C1. The vicilins from pea as well as vicilins from adzuki bean (Vigna angularis), garden bean (Phaseolus vulgaris), black-eyed pea (Vigna unguiculata), and mung bean (Vigna radiata), storage proteins that are homologous to the beta subunit of soybean beta-conglycinin, were not degraded by protease C1. Degradation of soybean beta-conglycinin involves a sequential attack of the alpha subunit at multiple sites, culminating in the formation of a stable intermediate of 53.5 kD and a final product of 48.0 kD. The cleavage sites resulting in this formation of the intermediates and final product were determined by N-terminal analysis. These were compared to the known amino acid sequences of the three beta-conglycinin subunits. Results showed these two polypeptides to be generated by proteolysis of the alpha subunit at regions bearing long strings of acidic amino acid residues.
Full Text
The Full Text of this article is available as a PDF (2.4M).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Bown D, Ellis TH, Gatehouse JA. The sequence of a gene encoding convicilin from pea (Pisum sativum L.) shows that convicilin differs from vicilin by an insertion near the N-terminus. Biochem J. 1988 May 1;251(3):717–726.[PMC free article] [PubMed] [Google Scholar]
- Croy RR, Gatehouse JA, Tyler M, Boulter D. The purification and characterization of a third storage protein (convicilin) from the seeds of pea (Pisum sativum L.). Biochem J. 1980 Nov 1;191(2):509–516.[PMC free article] [PubMed] [Google Scholar]
- Doyle JJ, Schuler MA, Godette WD, Zenger V, Beachy RN, Slightom JL. The glycosylated seed storage proteins of Glycine max and Phaseolus vulgaris. Structural homologies of genes and proteins. J Biol Chem. 1986 Jul 15;261(20):9228–9238. [PubMed] [Google Scholar]
- Garnier J, Osguthorpe DJ, Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. [PubMed] [Google Scholar]
- Gatehouse JA, Bown D, Gilroy J, Levasseur M, Castleton J, Ellis TH. Two genes encoding 'minor' legumin polypeptides in pea (Pisum sativum L.). Characterization and complete sequence of the LegJ gene. Biochem J. 1988 Feb 15;250(1):15–24.[PMC free article] [PubMed] [Google Scholar]
- Gatehouse JA, Lycett GW, Delauney AJ, Croy RR, Boulter D. Sequence specificity of the post-translational proteolytic cleavage of vicilin, a seed storage protein of pea (Pisum sativum L.). Biochem J. 1983 May 15;212(2):427–432.[PMC free article] [PubMed] [Google Scholar]
- Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed] [Google Scholar]
- Nielsen NC, Dickinson CD, Cho TJ, Thanh VH, Scallon BJ, Fischer RL, Sims TL, Drews GN, Goldberg RB. Characterization of the glycinin gene family in soybean. Plant Cell. 1989 Mar;1(3):313–328.[PMC free article] [PubMed] [Google Scholar]
- Prakash V, Rao MS. Physicochemical properties of oilseed proteins. CRC Crit Rev Biochem. 1986;20(3):265–363. [PubMed] [Google Scholar]
- Qi X, Wilson KA, Tan-Wilson AL. Characterization of the Major Protease Involved in the Soybean beta-Conglycinin Storage Protein Mobilization. Plant Physiol. 1992 Jun;99(2):725–733.[PMC free article] [PubMed] [Google Scholar]
- Sebastiani FL, Farrell LB, Schuler MA, Beachy RN. Complete sequence of a cDNA of alpha subunit of soybean beta-conglycinin. Plant Mol Biol. 1990 Jul;15(1):197–201. [PubMed] [Google Scholar]
- Thanh VH, Shibasaki K. Beta-conglycinin from soybean proteins. Isolation and immunological and physicochemical properties of the monomeric forms. Biochim Biophys Acta. 1977 Feb 22;490(2):370–384. [PubMed] [Google Scholar]
Abstract
Protease C1, an enzyme from soybean (Glycine max [L.] Merrill cv Amsoy 71) seedling cotyledons, was previously determined to be the enzyme responsible for the initial degradation of the alpha' and alpha subunits, but not the beta subunit, of beta-conglycinin storage protein. The sizes of the proteolytic products generated by the action of protease C1 suggest that the cleavage sites on the alpha' and alpha subunits of beta-conglycinin may be located in their N-terminal domain, which is not found in the beta subunit of beta-conglycinin. To check this hypothesis, storage proteins from other plant species that are homologous to either the alpha'/alpha or the beta subunit of beta-conglycinin were tested as substrates. As expected, the convicilin from pea (Pisum sativum), a protein homologous to the alpha' and alpha subunits of beta-conglycinin, was digested by protease C1. The vicilins from pea as well as vicilins from adzuki bean (Vigna angularis), garden bean (Phaseolus vulgaris), black-eyed pea (Vigna unguiculata), and mung bean (Vigna radiata), storage proteins that are homologous to the beta subunit of soybean beta-conglycinin, were not degraded by protease C1. Degradation of soybean beta-conglycinin involves a sequential attack of the alpha subunit at multiple sites, culminating in the formation of a stable intermediate of 53.5 kD and a final product of 48.0 kD. The cleavage sites resulting in this formation of the intermediates and final product were determined by N-terminal analysis. These were compared to the known amino acid sequences of the three beta-conglycinin subunits. Results showed these two polypeptides to be generated by proteolysis of the alpha subunit at regions bearing long strings of acidic amino acid residues.