Proteome analysis of hairy root from Panax ginseng C.A. Meyer using peptide fingerprinting, internal sequencing and expressed sequence tag data.
As an initial step to the comprehensive proteomic analysis of Panax ginseng C. A. Meyer, protein mixtures extracted from the cultured hairy root of Panax ginseng were separated by two-dimensional polyacrylamide gel electrophoresis (2-DE). The protein spots were analyzed and identified by peptide finger printing and internal amino acid sequencing by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) and electrospray ionization quadrupole-time of flight mass spectrometry (ESI Q-TOF MS), respectively. More than 300 protein spots were detected on silver stained two-dimensional (2-D) gels using pH 3-10, 4-7, and 4.5-5.5 gradients. Major protein spots (159) were analyzed by peptide fingerprinting or de novo sequencing and the functions of 91 of these proteins were identified. Protein identification was achieved using the expressed sequence tag (EST) database from Panax ginseng and the protein database of plants like Arabidopsis thaliana and Oryza sativa. However, peptide mass fingerprinting by MALDI-TOF MS alone was insufficient for protein identification because of the lack of a genome database for Panax ginseng. Only 17 of the 159 protein spots were verified by peptide mass fingerprinting using MALDI-TOF MS whereas 87 out of 102 protein spots, which included 13 of the 17 proteins identified by MALDI-TOF MS, were identified by internal amino acid sequencing using tandem mass spectrometry analysis by ESI Q-TOF MS. When the internal amino acid sequences were used as identification markers, the identification rate exceeded 85.3%, suggesting that a combination of internal sequencing and EST data analysis was an efficient identification method for proteome analysis of plants having incomplete genome data like ginseng. The 2-D patterns of the main root and leaves of Panax ginseng differed from that of the cultured hairy root, suggesting that some proteins are exclusively expressed by different tissues for specific cellular functions. Proteome analysis will undoubtedly be helpful for understanding the physiology of Panax ginseng.
Kim
Seung Il
SI
Proteome Analysis Team, Korea Basic Science Institute, 52 Yeoeun-Dong, Yusung-Ku, Daejon 305-333, Republic of Korea.
Kim
Jin Young
JY
Kim
Eun A
EA
Kwon
Kyung-Hoon
KH
Kim
Kyung-Wook
KW
Cho
Kun
K
Lee
Jeong Hwa
JH
Nam
Myung Hee
MH
Yang
Deok-Chun
DC
Yoo
Jong Shin
JS
Park
Young Mok
YM
D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D020224:Expressed Sequence Tags; D005894:Panax / Q000737:chemistry*; D010449:Peptide Mapping / Q000379:methods*; D010940:Plant Proteins / Q000032:analysis* / Q000737:chemistry*; D018517:Plant Roots / Q000737:chemistry; D020543:Proteome / Q000032:analysis* / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization
D010940:Plant Proteins; D020543:Proteome