Efficient peptide mapping and its application to identify embryo proteins in rice proteome analysis.
Using direct N-terminal analysis, only 31 N-terminally unblocked proteins out of 100 rice embryo proteins could be identified. To obtain protein sequence information for the remaining 69 blocked proteins, we developed a simple, efficient and rapid method. Using this method, we determined the peptide maps of 20 proteins per day in 10 pmol amounts. Applying this method to rice proteome analysis, we determined the internal sequences of all 69 blocked proteins. A total of 28 proteins out of 100 analyzed showed sequence similarity to the proteins with known functions in the SWISS-PROT and NCBI databases. Alternatively, we also used peptide mass fingerprinting determined by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) to identify the rice proteins separated by two-dimensional electrophoresis (2-DE). Although peptide-mass fingerprinting is a high-throughput method, we could not easily identify all the rice proteins or genes by this method, because the complete database information on rice, is not yet available and many proteins are post-translationally modified. Therefore, at present, the improved peptide mapping method as we report here is considered to be very useful in rice proteome analysis, especially for blocked proteins.
Woo
Sun-Hee
SH
Yokohama City University, Kihara Institute for Biological Research, Totsuka, Yokohama, Japan.
Fukuda
Masafumi
M
Islam
Nazrul
N
Takaoka
Motoko
M
Kawasaki
Hiroshi
H
Hirano
Hisashi
H
D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D012275:Oryza / Q000737:chemistry*; D010449:Peptide Mapping / Q000379:methods*; D010940:Plant Proteins / Q000032:analysis*; D020543:Proteome / Q000032:analysis; D012639:Seeds / Q000737:chemistry*; D020539:Sequence Analysis, Protein / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D010940:Plant Proteins; D020543:Proteome