Proteomic Approach of the Protein Profiles during Seed Maturation in Common Buckwheat (Fagopyrum esculentum Moench.)

Min-Hwa Park,Dong-Hoon Shin,Myoung-Hae Han,Young-Ho Yun,Jeong-Sook Bae,Yun-Sang Lee,Keun-Yook Chung,Moon-Soon Lee,Sun-Hee Woo 한국자원식물학회 2009 한국자원식물학회지 Vol.22 No.3

Single seeds of common buckwheat cultivar Suwon No. 1 when subjected to SDS-PAGE revealed very high polymorphism. High variation existed for protein or protein subunits with molecular weight 54-47kDa, 45-25kDa and 16-11kDa. The electrophoregram showed variation for globulin as well as other protein fractions. About 300 proteins were separated by two-dimensional electrophoresis in common buckwheat (Fagopyrum esculentum Moench.) seed. Seed maturation is a dynamic and temporally regulated phase of seed development that determines the composition of storage proteins reserves in mature seeds. Buckwheat seeds from 5, 10, 15, 20, and 25 days after pollination and matured stage were used for the analysis. This led to the establishment of high-resolution proteome reference maps, expression profiles of 48 spots. It was identified 48 proteins from MALDI-TOF/MS analysis of wild buckwheat seed storage proteins. The 48 proteins were found identical or similar to those of proteins reported in buckwheat and other plants; it is belonging to 9 major functional categories including seed storage proteins, stress/defense response, protein synthesis, photosynthesis, allergy proteins, amino acid, enzyme, metabolism, and miscellaneous. It appears that the major allergenic storage protein separated played the important role in buckwheat breeding and biochemical characterization.

Effects of Reduced Prolamin on Seed Storage Protein Composition and the Nutritional Quality of Rice

Kim, Hyun-Jung,Lee, Jong-Yeol,Yoon, Ung-Han,Lim, Sun-Hyung,Kim, Young-Mi Molecular Diversity Preservation International (MD 2013 INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES Vol.14 No.8

<P>Rice seed storage proteins accumulate in two types of protein body (PB-I and PB-II) that are nutrient sources for animals. PB-I is indigestible and negatively affects rice protein quality. To improve the nutritional value of rice seeds we are aiming to engineer the composition and accumulation of endogenous seed storage proteins. In this study we generated transgenic rice plants in which 13 kD prolamin genes were suppressed by RNA interference (13 kD pro-RNAi). Analysis based on qRT-PCR confirmed that the targeted 13 kD prolamins were markedly suppressed, and were compensated for by an increase in other storage proteins including 10 kD prolamin, glutelins, and chaperone proteins. The storage protein profiles further revealed that the levels of 13 kD prolamins were significantly reduced, while that of the glutelin precursor was slightly increased and the remaining storage proteins did not change. Amino acid analysis showed that the reduction of 13 kD prolamins resulted in a 28% increase in the lysine content relative to the wild type, indicating that the 13 kD pro-RNAi rice seeds are more nutritious. Furthermore, a reduction in the levels of 13 kD prolamins resulted in abnormal formation of PB-I, which was small and had no lamellar structure. These results suggest that alteration of prolamins can contribute to improving the nutritional quality of rice.</P>

Variations in Seed Storage Protein among Different Colored Soybean Varieties

Kim, Sun-Lim,Yun, Hong-Tae,Moon, Jung-Kyung,Park, Keum-Yong,Lee, Yeong-Ho,Ryu, Yong-Hwan The Korean Society of Crop Science 2004 Korean journal of crop science Vol.49 No.2

This study was carried out to know the variation of soybean seed proteins, 11S and 7S globulins, and their amino acid compositions among different colored soybean varieties, 'Danbaegkong' (yellow), 'Pureunkong' (green) 'Jinyulkong' (brown), and 'Geoumjeongkong l' (black). Soybean seed proteins showed a wide range in molecular size, but the electrophoresis patterns of total seed protein subunits showed a similarity among different colored soybean varieties. Amino acid compositions of total seed proteins were similar for all soybean varieties tested. However, soybean varieties showed low composition rates in sulfur containing amino acids. The composition rates of cysteine and methionine in the 11S globulins were higher than those of total seed proteins and 7S globulins. Glutamic acid and glycine were higher in the 11S and 7S globulins than those of total seed proteins. However, the levels of methionine and phenylalanine are high in the 11S globulins, but those of valine and lysin are slightly lower than the 7S globulins. By using HPLC, we tried to analyse the soybean seed proteins. The 11S globulin was composed of 10 major peaks whereas the 7S globulin was composed of 4 major peaks. The composition rates of 11S related proteins have a tendency to increasing during the maturing whereas those of 7S related proteins have a tendency to decreasing. Composition rates of each peaks among different colored soybean varieties suggested that soybean seed proteins are varied, although they showed similarity in the electrophoresis patterns, and understanding of this characteristics is important for the utilization of soybeans.

콩 유전자원의 종실 저장단백질 변이 및 유용자원 발굴

허건 ( Gun Hur ),장성진 ( Seong Jin Jang ),이윤호 ( Yun Ho Lee ),송항림 ( Hang Lin Song ),김성진 ( Seong Jin Kim ),정헌상 ( Heon Sang Jeong ),조용구 ( Yong Gu Cho ),김홍식 ( Hong Sig Kim ) 한국콩연구회 2008 韓國콩硏究會誌 Vol.25 No.1

한국야생콩 200종, 한국재래콩 300종 및 도입유전자원 398종을 포함한 총 898종에 대하여 SDS-PAGE에 의한 종실 단백질의 변이를 구명하고 변이자원을 발굴하여 고품질 콩 품종개발의 기초자료로 이용코자 수행하였다. 1. 7S함량은 평균 45.85%로서 7.16-66.24%의 범위였고, 변이자원은 α` subunit 결실 6종, α subunit 결실 1종, α` 및 α subunit 동시 결실 2종, β subunit 결실 6종, α 및 β subunit 동시저하 1종의 총 16종이 발굴되었다. 2. 11S함량은 평균 54.16%로서 33.76-89.67%의 범위였고, 변이자원은 A3 subunit 결실 6종, Acidic 저하 3종, Basic 저하 5종, Acidic 및 Basic 동시저하 1종의 총 15종이 발굴되었다. 3. 11S/7S 비율은 평균 1.18로서 0.5-8.68의 범위였으며, 도입유전자원의 변이가 가장 컸고, 다음으로 한국 야생콩이 컸으며 한국 재래콩이 작았다. 11S/7S 비율이 높은 자원으로는 CS1733, CS79, CS1228, CS1230 및 CS667의 5종이 발굴되었다. 4. Lipoxygenase 결실 변이자원은 11종, Kunitz trypsin inhibitor 결실 변이자원은 12종이 발굴되었다. To identify soybean genetic resources with specific seed proteins, a total of 898 soybean resources including 200 Korean wild soybeans, 300 korean local soybeans and 398 introduced soybeans were screened to evaluate variations of seed storage protein by SDS-PAGE. 7S concentrations of seed storage protein ranged from 7.16% to 66.24% with a mean of 45.85%. Sixteen variants including six lacking of α` subunit, one lacking of α subunit, two lacking of both α` and α subunits, six lacking of β subunit and one low level of both α and β subunits were detected for 7S seed storage protein. 11S concentrations of seed storage protein ranged from 33.76% to 89.67% with a mean of 54.16%. Fifteen variants including six lacking of A3 subunit, three low level of acidic group, five low level of basic group were detected for 11S seed storage protein. 11S/7S ratio of seed storage protein ranged from 0.5 to 8.68 with a mean of 1.18. Variation of 11S/7S ratio was largest in introduced soybeans, medium in Korean wild soybeans, and smallest in Korean local soybeans among three soybean groups. Of 898 soybean germplasms, CS1733, CS79, CS1228, CS1230 and CS667 were extremely high in 11S/7S ratio of seed storage protein. Eleven and twelve variants were detected for lacking lipoxygenase and for lacking kunitz trypsin inhibitor, respectively.

A Systematic Proteome Study of Seed Storage Proteins from Two Soybean Genotypes

Seong-Woo Cho,Soo-Jeong Kwon,Swapan Kumar Roy,Hong-Sig Kim,Chul-Won Lee,Sun Hee Woo 한국작물학회 2014 Korean journal of crop science Vol.59 No.3

Soybean seed is a good source of plant protein in human consumables such as baby formula and protein concentrate. The seeds contain an abundance of storage proteins, namely β-conglycin and glycinin that account for ~ 70-80% of the total seed protein content. Proteome profiling has been proved to be an efficient way that can help us to investigate the seed storage proteins. In the present study, the seeds were removed from the pods and the cotylendonary tissues were separated from the testa for proteome analysis in order to investigate the seed storage proteins. A systematic proteome profiling was conducted through one-dimensional gel electrophoresis followed by MALDI-TOF-TOF mass spectrometry in the seeds (cotyledonary tissue) of soybean genotypes. Two dimensional gels stained with CBB, a total of 10 proteins were identified and analyzed using MASCOT search engine according to the similarity of sequences with previously characterized proteins along with the UniProt database. A total of ten proteins such as glycinin Gy4 precursor, glycinin G3 precursor, glycinin G1 precursor, glycinin chain A2B1a precursor, glycinin chain A2B1a precursor were identified in our investigation. However, the glycinin subunit may be considered to play important roles in soybean breeding and biochemical characterization. In addition, the improved technique will be useful to dissect the genetic control of glycinin expression in soybean.

콩의 7S α′- subunit 단백질의 유전

성미경,박정수,김경록,황교진,정종일 경상대학교 농업생명과학연구원 2009 농업생명과학연구 Vol.43 No.5

Soybean is an important sources of plant proteins for human and animal nutrition. The use of soybean proteins has been expanded in the food industry due to their excellent nutritional benefits. But, Soybeans contain allergenic proteins that cause allergies to sensitive individuals. β-conglycinin(7S globulin) and glycinin(11S globulin) are the major components of storage protein in soybean. β-conglycinin consists of three subunits, α′, α, β and exhibits poorer nutritional and food processing properties than glycinin. There is a great deal of interest in the development of soybean lines with reduced amounts of β-conglycinin. The objective of this study was to determine the inheritance of α′-subunit protein in 7S globulin. F2 population was developed from the cross of "Jinpumkong2ho"(α′-subunit presence) and PI506876(α′-subunit absence) parent. Total 98 of F2 seeds were obtained and analyzed for the segregation of α′-subunit protein by SDS-PAGE. Among 98 F2 seeds, 70 F2 seeds showed α′-subunit protein and 28 F2 seeds did not show α′-subunit protein. The segregation ratios of 3 : 1 for presence and absence of α′-subunit protein were observed(χ2=0.667, P=0.414). These data indicate that presence and absence of α′-subunit protein is controlled by a single major gene and might be useful for strain selection of 7S protein reduced soybean. 인간과 가축의 영양을 위한 식물성 단백질의 주요 공급원은 콩이며 콩 단백질은 영양 및 기능성면에서 우수하여 소비가 점차 증가하고 있다. 그러나 콩 단백질에는 알러지를 일으키고 영양가치를 떨어뜨리는 성분도 포함되어져 있다. 7S 및 11S 글로블린은 콩 저장단백질의 대부분을 차지하며 7S는 영양가치가 떨어지고 7S의 함량을 줄어든 콩 계통 육성에 대한 관심이 높아지고 있다. 7S 성분중의 하나인 α′-subunit의 유전양상을 파악하기 위하여 진품콩2호와 PI506876의 교배로부터 98개의 F2 종자가 얻어졌다. SDS-PAGE로 각각의 종자를 분석한 결과 α′-subunit을 가진 종자가 70개였고 결핍된 종자가 28개였다. 이러한 유전양상은 단인자 유전원칙 (χ2 = 0.667, P=0.414)과 일치하여 콩 종자에서 7S의 α′-subunit 단백질은 한 개의 유전자에 의해서 좌우되었다. 이 결과는 7S 단백질 함량이 줄어든 콩 계통 선발에 유용하게 활용될 것으로 기대된다.

A Systematic Proteome Study of Seed Storage Proteins from Two Soybean Genotypes

Cho, Seong-Woo,Kwon, Soo-Jeong,Roy, Swapan Kumar,Kim, Hong-Sig,Lee, Chul-Won,Woo, Sun Hee The Korean Society of Crop Science 2014 한국작물학회지 Vol.59 No.3

Soybean seed is a good source of plant protein in human consumables such as baby formula and protein concentrate. The seeds contain an abundance of storage proteins, namely ${\beta}$-conglycin and glycinin that account for ~ 70-80% of the total seed protein content. Proteome profiling has been proved to be an efficient way that can help us to investigate the seed storage proteins. In the present study, the seeds were removed from the pods and the cotylendonary tissues were separated from the testa for proteome analysis in order to investigate the seed storage proteins. A systematic proteome profiling was conducted through one-dimensional gel electrophoresis followed by MALDI-TOF-TOF mass spectrometry in the seeds (cotyledonary tissue) of soybean genotypes. Two dimensional gels stained with CBB, a total of 10 proteins were identified and analyzed using MASCOT search engine according to the similarity of sequences with previously characterized proteins along with the UniProt database. A total of ten proteins such as glycinin Gy4 precursor, glycinin G3 precursor, glycinin G1 precursor, glycinin chain A2B1a precursor, glycinin chain A2B1a precursor were identified in our investigation. However, the glycinin subunit may be considered to play important roles in soybean breeding and biochemical characterization. In addition, the improved technique will be useful to dissect the genetic control of glycinin expression in soybean.

Variations in Seed Storage Protein among Different Colored Soybean Varieties

Sun-Lim Kim,Hong-Tae Yun,Jung-Kyung Moon,Keum-Yong Park,Yeong-Ho Lee,Yong-Hwan Ryu 韓國作物學會 2004 Korean journal of crop science Vol.49 No.2

This study was carried out to know the variation of soybean seed proteins, 11S and 7S globulins, and their amino acid compositions among different colored soybean varieties, 'Danbaegkong' (yellow), 'Pureunkong' (green) 'Jinyulkong' (brown), and 'Geoumjeongkong l' (black). Soybean seed proteins showed a wide range in molecular size, but the electrophoresis patterns of total seed protein subunits showed a similarity among different colored soybean varieties. Amino acid compositions of total seed proteins were similar for all soybean varieties tested. However, soybean varieties showed low composition rates in sulfur containing amino acids. The composition rates of cysteine and methionine in the 11S globulins were higher than those of total seed proteins and 7S globulins. Glutamic acid and glycine were higher in the 11S and 7S globulins than those of total seed proteins. However, the levels of methionine and phenylalanine are high in the 11S globulins, but those of valine and lysin are slightly lower than the 7S globulins. By using HPLC, we tried to analyse the soybean seed proteins. The 11S globulin was composed of 10 major peaks whereas the 7S globulin was composed of 4 major peaks. The composition rates of 11S related proteins have a tendency to increasing during the maturing whereas those of 7S related proteins have a tendency to decreasing. Composition rates of each peaks among different colored soybean varieties suggested that soybean seed proteins are varied, although they showed similarity in the electrophoresis patterns, and understanding of this characteristics is important for the utilization of soybeans.

Proteomic Analysis of Seed Storage Proteins in Low Allergenic Soybean Accession

( Sung Cheol Koo ),( Dong Won Bae ),( Jun Su Seo ),( Kyoung Mi Park ),( Man Soo Choi ),( Seok Hyeon Kim ),( Sang In Shim ),( Kyeong Moon Kim ),( Jong Il Chung ),( Min Chul Kim ) 한국응용생명화학회 2011 Applied Biological Chemistry (Appl Biol Chem) Vol.54 No.3

Soybean (Glycine max L. Merr.) is a major source of protein for human and animal nutrients. However, soybean seed also contains a number of allergenic proteins, which limit its extensive usage in the food industry. P34 (Gly m Bd 30K), a low-leveled but highly conserved seed storage protein, has been described as a major allergen in soybean seed. Recently, two low P34 soybean accessions, PI603570A and PI567476, were identified by extensive screening of soybean germplasm. We have surveyed spatiotemporal expression pattern of P34 protein in low P34 accession, PI567476, and cultivated soybean accession, Clark, using antibody against P34. Immunoblot analysis showed not only seed specific expression pattern of P34, but also significantly reduced expression of P34 in PI567476 compared to Clark. Additionally, we performed proteomic analysis to compare protein compositions between PI567476 and Clark accessions. Although the overall distribution pattern of the protein spots was similar in both soybean accessions, 19 protein spots showing different intensity between both accessions were detected and analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS). PI567476 accession exhibited altered expression of several seed storage proteins including allergens and seed maturation-related proteins. The detailed analysis for composition of seed storage proteins of the PI567476 accession provides valuable information in use of this soybean germplasm for breeding of low allergenic elite soybean lines, which is expected to enhance food safety and make soy products available to allergen-sensitive individuals.

A TMT-based quantitative proteomic analysis provides insights into the protein changes in the seeds of high- and low- protein content soybean cultivars

민철우,GUPTARAVI,NGUYEN VAN TRUONG,배진우,고종민,이병원,김선태 한국식물생명공학회 2020 JOURNAL OF PLANT BIOTECHNOLOGY Vol.47 No.3

The presence of high amounts of seed storage proteins (SSPs) improves the overall quality of soybean seeds. However, these SSPs pose a major limitation due to their high abundance in soybean seeds. Although various technical advancements including mass-spectrometry and bioinformatics resources were reported, only limited information has been derived to date on soybean seeds at proteome level. Here, we applied a tandem mass tags (TMT)-based quantitative proteomic analysis to identify the significantly modulated proteins in the seeds of two soybean cultivars showing varying protein contents. This approach led to the identification of 5,678 proteins of which 13 and 1,133 proteins showed significant changes in Daewon (low-protein content cultivar) and Saedanbaek (high-protein content cultivar) respectively. Functional annotation revealed that proteins with increased abundance in Saedanbaek were mainly associated with the amino acid and protein metabolism involved in protein synthesis, folding, targeting, and degradation. Taken together, the results presented here provide a pipeline for soybean seed proteome analysis and contribute a better understanding of proteomic changes that may lead to alteration in the protein contents in soybean seeds.