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7S and 11S seed storage globulins: evolution, proteolysis, allergenicity


Author: Cherdivară Ala
Degree:doctor of biology
Speciality: 03.00.04 - Biochemistry
Year:2020
Scientific adviser: Andrei Şutov
doctor habilitat, associate professor (docent)
Institution: Moldova State University

Status

Term of presenting of the thesis 4 December, 2020

Abstract

Adobe PDF document4.59 Mb / in romanian

Thesis

CZU 577.1./.2(043)

Adobe PDF document 16.00 Mb / in romanian
132 pages


Keywords

seed storage globulins, legumin, vicilin, germin, oxalate decarboxylase, evolution, protein primary and higher order structures, proteolysis mechanisms, limited proteolysis, proteolysis kinetics, allergenicity, ginkgo, peanut

Summary

The work contains 116 pages of the text, 64 figures, 14 tables and the list of references that includes 120 sources. The obtained results were published in 18 scientific papers (9 articles and 9 reports at conferences).

Aim of the study: Investigation of the regularities of formation during evolution of the conserved structure of seed storage globulins, establishment of the mechanism of their deep proteolytic degradation and elucidation of the prospects for the use of limited proteolysis of storage globulins to reduce the level of their allergenicity. The objectives of the study. 1) To analyze phylogenetic relationships between sequences of seed storage globulins and related proteins of prokaryotes and eukaryotes. 2) To establish successive stages in the evolution of the structure of storage globulins that determine their function. 3) To study the kinetics of proteolysis of the storage 11S globulin of ginkgo Ginkgo biloba, one of the most ancient representatives of spermatophytes, and to describe the mechanism of regulation of deep proteolytic degradation of 11S globulins. 4) To study the successive reactions of limited proteolysis of 11S and 7S globulins from peanut Arachis hypogaea, whose seeds are known for a high level of allergenicity. 5) To establish structural changes in 11S and 7S globulins during proteolysis and assess the possibility of reducing the allergenicity of peanut storage globulins by limited proteolysis.

Scientific novelty and originality of the work. It was established for the first time that bacterial oxalate decarboxylases represent statistically reliable most ancient precursors of seed storage globulins. It was demonstrated that proteins from the green algae Klebsormidium nitens represent the most ancient plant precursors of seed storage globulins. It was shown that the primary and higher order structures of 11S globulins reflect the most ancient features of seed storage globulins. It was demonstrated that the limited proteolysis of 11S globulin from ginkgo seeds initiates massive degradation of the protein via one-by-one mechanism. Based on the analysis of crystal structures of 11S globulins, the regulatory role of the limited proteolysis in the massive degradation of these proteins was shown. Regularities of limited proteolysis of 11S and 7S globulins from pumpkin seeds were studied.

It was demonstrated that limited proteolysis leads to the removal of the primary structure regions of peanut 11S and 7S globulins, which determine their allergenicity. The obtained result, which contributes to the solution of an important scientific problem, is to establish the origin of seed storage globulins from bacterial oxalate decarboxylases and the regulatory role of limited proteolysis in their deep degradation. It has been shown that limited proteolysis of seed storage globulins leads to the removal of antigenic determinants (IgE epitopes) from their primary structures, which is promising from the point of view of increasing the safety of food products containing seed proteins.

Theoretical significance of the work. The origin of seed storage globulins from bacterial oxalate decarboxylases, rather than from Amoebozoa spherulins as assumed previously, was shown statistically faithfully. The role of limited proteolysis in the regulation of massive proteolytic degradation of 11S globulin from ginkgo, one of the oldest representatives of seed plants, has been established. The results obtained suggest a common mechanism for the regulation of proteolysis of storage proteins characteristic of the entire family of 11S seed globulins. Practical significance of the work. The availability of the limited proteolysis as a means of a significant decrease of the allergenicity level of storage globulins from pumpkin seeds, as well as from seeds of other plants, were shown.

Implementation of scientific results. The results presented in the paper represent scientific-didactic material for the courses of biochemistry and proteomics.