Signal-anchor domains of proteins of the outer membrane of mitochondria: structural and functional characteristics.
We have studied the topogenesis of a class of mitochondrial outer membrane proteins that expose a hydrophilic domain to the cytosol and are anchored to the membrane by a single transmembrane domain in the N-terminal region. To determine the role of these latter sequences in the targeting and insertion of such proteins we took two approaches. First, a functional complementation assay was used to define the structural elements that together with the anchor domain make up the topogenic signal. Moderate hydrophobicity of the transmembrane domain was found to be the most important requirement. Variants with a scrambled sequence of the membrane-spanning segment were only partially functional suggesting that specificity in the amino acid sequence is also of considerable importance. A net positive charge at both flanking regions of the transmembrane domain contributes to the efficiency of targeting and membrane integration but is not an essential structural feature of this signal. Second, chimeras of Tom20, Tom70, and OM45 were generated that contained the cytosolic domain of Tom20 or Tom70 and the anchor domain of one of the other members of the class. These hybrid proteins were able to rescue the growth of cells lacking Tom20 or Tom70. Thus, anchor domains of outer membrane proteins are functionally interchangeable. They play only a minor role in the specific function of these proteins, but have a decisive role in topogenic signaling.
Waizenegger
Thomas
T
Institut für Physiologische Chemie der Universität München, Butenandtstrasse 5, Haus B, D-81377 Munich, Germany.
Stan
Tincuta
T
Neupert
Walter
W
Rapaport
Doron
D
D000595:Amino Acid Sequence; D005656:Fungal Proteins / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D007425:Intracellular Membranes / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D008928:Mitochondria / Q000648:ultrastructure; D024101:Mitochondrial Proteins / Q000737:chemistry*; D021382:Protein Sorting Signals; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins
D005656:Fungal Proteins; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins