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Primary structure of a novel subunit in ba3-cytochrome oxidase from Thermus thermophilus

Published online by Cambridge University Press:  15 December 2000

TEWFIK SOULIMANE
Affiliation:
Rheinisch-Westfälische Technische Hochschule Aachen, Institut für Biochemie, Pauwelsstraße 30, D-52057 Aachen, Germany
MANUEL E. THAN
Affiliation:
Max-Planck Institut für Biochemie, Am Klopferspitz 18a, D-82152 Planegg-Martinsried, Germany
MANFRED DEWOR
Affiliation:
Rheinisch-Westfälische Technische Hochschule Aachen, Institut für Biochemie, Pauwelsstraße 30, D-52057 Aachen, Germany
ROBERT HUBER
Affiliation:
Max-Planck Institut für Biochemie, Am Klopferspitz 18a, D-82152 Planegg-Martinsried, Germany
GERHARD BUSE
Affiliation:
Rheinisch-Westfälische Technische Hochschule Aachen, Institut für Biochemie, Pauwelsstraße 30, D-52057 Aachen, Germany
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Abstract

The ba3-type cytochrome c oxidase from Thermus thermophilus is known as a two subunit enzyme. Deduced from the crystal structure of this enzyme, we discovered the presence of an additional transmembrane helix “subunit IIa” spanning the membrane. The hydrophobic N-terminally blocked protein was isolated in high yield using high-performance liquid chromatography. Its complete amino acid sequence was determined by a combination of automated Edman degradation of both the deformylated and the cyanogen bromide cleaved protein and automated C-terminal sequencing of the native protein.

The molecular mass of 3,794 Da as determined by MALDI-MS and by ESI requires the N-terminal methionine to be formylated and is in good agreement with the value calculated from the formylmethionine containing sequence (3,766.5 Da + 28 Da = 3,794.5 Da). This subunit consits of 34 residues forming one helix across the membrane (Lys5–Ala34), which corresponds in space to the first transmembrane helix of subunit II of the cytochrome c oxidases from Paracoccus denitrificans and bovine heart, however, with opposite polarity. It is 35% identical to subunit IV of the ba3-cytochrome oxidase from Natronobacterium pharaonis.

The open reading frame encoding this new subunit IIa (cbaD) is located upstream of cbaB in the same operon as the genes for subunit I (cbaA) and subunit II (cbaB).

Type
Research Article
Copyright
2000 The Protein Society

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