Systematic cyanobacterial membrane proteome analysis by combining acid hydrolysis and digestive enzymes with nano-liquid chromatography-Fourier transform mass spectrometry.
The identification of membrane proteins is currently under-represented since the trans-membrane domains of membrane proteins have a hydrophobic property. Membrane proteins have mainly been analyzed by cleaving and identifying exposed hydrophilic domains. We developed the membrane proteomics method for targeting integral membrane proteins by the following sequential process: in-solution acid hydrolysis, reverse phase chromatographic separation, trypsin or chymotrypsin digestion and nano-liquid chromatography-Fourier transform mass spectrometry. When we employed total membrane proteins of Synechocystis sp. PCC 6803, 155 integral membrane proteins out of a predictable 706 were identified in a single application, corresponding to 22% of a genome. The combined methods of acid hydrolysis-trypsin (AT) and acid hydrolysis-chymotrypsin (AC) identified both hydrophilic and hydrophobic domains of integral membrane proteins, respectively. The systematic approach revealed a more concrete data in mapping the repertoire of cyanobacterial membrane and membrane-linked proteome.
Kwon
Joseph
J
Gwangju Center, Korea Basic Science Institute, Gwangju 500-757, South Korea.
Oh
Jeehyun
J
Park
Chiyoul
C
Cho
Kun
K
Kim
Seung Il
SI
Kim
Soohyun
S
Lee
Sunghoon
S
Bhak
Jong
J
Norling
Birgitta
B
Choi
Jong-Soon
JS
D001426:Bacterial Proteins / Q000032:analysis* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002918:Chymotrypsin / Q000378:metabolism; D003198:Computer Simulation; D005583:Fourier Analysis; D006868:Hydrolysis; D013058:Mass Spectrometry / Q000379:methods*; D008565:Membrane Proteins / Q000032:analysis* / Q000378:metabolism; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis / Q000378:metabolism; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D046939:Synechocystis / Q000737:chemistry*; D014357:Trypsin / Q000378:metabolism
D001426:Bacterial Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D020543:Proteome; D002918:Chymotrypsin; D014357:Trypsin