A simple search of TM segments in polytopic membrane protein using matrix-assisted laser desorption ionization time-of-flight mass spectrometry.
Using both high performance liquid chromatography (HPLC) and amino acid sequencing (AAS), we previously analyzed band 3 TM peptide-segments that make up the transmembrane protein structure. However, the HPLC/AAS combination method was highly time-consuming. Matrix-Assisted Laser Desorption Ionization Time-of-Flight (MALDI-TOF) mass spectrometry is used to obtain accurate molecular weight information for proteins/peptides simply and sensitively. We applied the MALDI-TOF mass spectrometry technique to search for TM segments in membrane proteins. In combination with trypsin cleavages after alkali treatments (pH12 or 13) and sample preparation using organic solvents for MALDI-TOF mass spectrometry, we determined the TM segments of band 3 and glycophorin A in erythrocyte membrane. The method can be applied to other polytopic membrane proteins in erythrocyte membrane.
Abe
Yoshito
Y
Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan. abe@phar.kyushu-u.ac.jp
Hamasaki
Tomohiro
T
Turusaki
Susumu
S
Takazaki
Shinya
S
Jin
Xiuri
X
Kang
Dongchon
D
Hamasaki
Naotaka
N
D000468:Alkalies / Q000378:metabolism; D000595:Amino Acid Sequence; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014357:Trypsin / Q000378:metabolism
D000468:Alkalies; D008565:Membrane Proteins; D014357:Trypsin