Identification of peptide fragments chemically cross-linked in cytochrome c oxidase from thermophilic Bacillus PS3.
In order to study steric arrangement of subunits in cytochrome c oxidase isolated from thermophilic Bacillus PS3, we developed a simple procedure including chemical cross-linking, two consecutive runs of electrophoresis, proteolytic digestion, and peptide sequencing for simultaneous identification of two cross-linked fragments. By this procedure, the cytochrome c domain of subunit 2 was found cross-linked to C-terminal region of subunit 1 including two hydrophobic transmembrane segments, suggesting that these two regions were located close each other. The present simple procedure might be applicable to proteins whose crystal structures are not revealed.
Kuge
S
S
Department of Biochemical Engineering and Science, Kyushu Institute of Technology, Fukuoka, Japan.
Noguchi
S
S
Sakamoto
J
J
Sone
N
N
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology*; D003432:Cross-Linking Reagents / Q000737:chemistry*; D003576:Electron Transport Complex IV / Q000737:chemistry* / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry*; D011804:Quinolines / Q000378:metabolism; D017421:Sequence Analysis; D012697:Serine Endopeptidases / Q000378:metabolism; D013388:Succinimides / Q000737:chemistry
D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D010446:Peptide Fragments; D011804:Quinolines; D013388:Succinimides; C000470:EEDQ; C018052:disuccinimidyl tartarate; C028722:ethylene glycolylbis(succinimidyl succinate); D003576:Electron Transport Complex IV; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; C011240:dithiobis(succinimidylpropionate)