Thylakoid membrane at altered metabolic state: challenging the forgotten realms of the proteome.
Analysis of the membrane integral proteome is mainly dependent on the ability of protein separation. Blue-native polyacrylamide gel electrophoresis (BN-PAGE) is a technique capable of efficient membrane protein separation, so far mainly applied to the mitochondrial oxidative phosphorylation machinery. Applying BN-PAGE to the thylakoid membranes after mild solubilization with digitonin we succeeded in displaying the response of the green algae Chlamydomonas reinhardtii to altered culture conditions. In addition, by peptide mass fingerprinting and matrix assisted laser desorption/ionization-mass spectrometry (MALDI-MS) extremely hydrophobic subunits of the photosystem complexes with 5-11 transmembrane helices were identified, which could not be accessed by in-gel digestion in previous studies.
Rexroth
Sascha
S
Department of Chemistry, Physical Biochemistry, Damstadt University of Technology, Petersenstrasse 22, D-64287 Darmstadt, Germany. srex@gmx.de
Meyer zu Tittingdorf
Jürgen M W
JM
Krause
Frank
F
Dencher
Norbert A
NA
Seelert
Holger
H
D000222:Adaptation, Physiological; D020418:Algal Proteins / Q000032:analysis / Q000302:isolation & purification / Q000528:radiation effects; D000460:Chlorophyta / Q000737:chemistry / Q000528:radiation effects; D004072:Digitonin; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation / Q000528:radiation effects; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000032:analysis / Q000302:isolation & purification / Q000528:radiation effects; D045332:Photosystem II Protein Complex / Q000032:analysis / Q000737:chemistry* / Q000528:radiation effects; D021122:Protein Subunits / Q000302:isolation & purification; D020543:Proteome / Q000032:analysis / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D020524:Thylakoids / Q000378:metabolism* / Q000528:radiation effects
D020418:Algal Proteins; D008565:Membrane Proteins; D045332:Photosystem II Protein Complex; D021122:Protein Subunits; D020543:Proteome; D004072:Digitonin