TY - JOUR
T1 - Subunit composition of Rhodothermus marinus respiratory complex I
AU - Pereira, Manuela Alexandra
AU - Coelho, Ana Maria
N1 - Batista, Ana P.; Franco, Catarina; Mendes, Marta;
PY - 2010/1/1
Y1 - 2010/1/1
N2 - The basic structural characterization of complex I is still not trivial due to its complexity, not only in the number of its protein constituents but especially because of the different properties of the several subunits. Bacterial complex I generally contains 14 subunits: 7 hydrophilic proteins located in the peripheral arm and 7 hydrophobic proteins present in the membrane arm. It is the identification of the hydrophobic proteins that makes the characterization of complex I, and of membrane proteins in general, very difficult. In this article, we report the identification of the subunits of complex I from Rhodothermus marinus. The original approach, presented here, combined several protein and peptides separation strategies (different reversed phase materials, high-performance liquid chromatography, and gel electrophoresis) with different identification methods (electrospray ionization-tandem mass spectrometry, matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry, and Edman degradation analysis) and represents a step forward in the characterization of membrane proteins that studies are still technically highly challenging. The combination of the different methodologies allowed the identification of complex I canonical subunits and also a possible novel subunit, namely a pterin-4 alpha-carbinolamine dehydratase (PCD). This was the first time that a PCD was suggested to be part of complex I, and its possible regulatory role is discussed.
AB - The basic structural characterization of complex I is still not trivial due to its complexity, not only in the number of its protein constituents but especially because of the different properties of the several subunits. Bacterial complex I generally contains 14 subunits: 7 hydrophilic proteins located in the peripheral arm and 7 hydrophobic proteins present in the membrane arm. It is the identification of the hydrophobic proteins that makes the characterization of complex I, and of membrane proteins in general, very difficult. In this article, we report the identification of the subunits of complex I from Rhodothermus marinus. The original approach, presented here, combined several protein and peptides separation strategies (different reversed phase materials, high-performance liquid chromatography, and gel electrophoresis) with different identification methods (electrospray ionization-tandem mass spectrometry, matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry, and Edman degradation analysis) and represents a step forward in the characterization of membrane proteins that studies are still technically highly challenging. The combination of the different methodologies allowed the identification of complex I canonical subunits and also a possible novel subunit, namely a pterin-4 alpha-carbinolamine dehydratase (PCD). This was the first time that a PCD was suggested to be part of complex I, and its possible regulatory role is discussed.
KW - UBIQUINONE OXIDOREDUCTASE
KW - NADH
KW - IDENTIFICATION
KW - PSEUDOMONAS-AERUGINOSA
KW - PEPTIDE MIXTURES
KW - NDH-1 COMPLEXES
KW - DESORPTION/IONIZATION-MASS-SPECTROMETRY
KW - MEMBRANE-PROTEINS
KW - PHENYLALANINE-HYDROXYLASE
KW - PTERIN-4A-CARBINOLAMINE DEHYDRATASE
U2 - 10.1016/j.ab.2010.07.038
DO - 10.1016/j.ab.2010.07.038
M3 - Article
VL - 407
SP - 104
EP - 110
JO - Analytical Biochemistry
JF - Analytical Biochemistry
SN - 0003-2697
IS - 1
ER -