Limits...
The TOM core complex: the general protein import pore of the outer membrane of mitochondria.

Ahting U, Thun C, Hegerl R, Typke D, Nargang FE, Neupert W, Nussberger S - J. Cell Biol. (1999)

Bottom Line: It forms a double ring structure that, in contrast to the holo complex, lacks the third density seen in the latter particles.Three-dimensional reconstruction by electron tomography exhibits two open pores traversing the complex with a diameter of approximately 2.1 nm and a height of approximately 7 nm.Tom40 is the key structural element of the TOM core complex.

View Article: PubMed Central - PubMed

Affiliation: Institut für Physiologische Chemie der Universität München, D-80336 München, Germany.

ABSTRACT
Translocation of nuclear-encoded preproteins across the outer membrane of mitochondria is mediated by the multicomponent transmembrane TOM complex. We have isolated the TOM core complex of Neurospora crassa by removing the receptors Tom70 and Tom20 from the isolated TOM holo complex by treatment with the detergent dodecyl maltoside. It consists of Tom40, Tom22, and the small Tom components, Tom6 and Tom7. This core complex was also purified directly from mitochondria after solubilization with dodecyl maltoside. The TOM core complex has the characteristics of the general insertion pore; it contains high-conductance channels and binds preprotein in a targeting sequence-dependent manner. It forms a double ring structure that, in contrast to the holo complex, lacks the third density seen in the latter particles. Three-dimensional reconstruction by electron tomography exhibits two open pores traversing the complex with a diameter of approximately 2.1 nm and a height of approximately 7 nm. Tom40 is the key structural element of the TOM core complex.

Show MeSH
EM and projection map of the TOM core complex. A, Survey view of negatively stained TOM core complex. The image was filtered to the first zero of the electron microscope contrast transfer function. Bar, 11 nm. B, Classification analysis of 1,598 TOM core complex particles. Using MSA, the data set was split into 20 classes. Classes 1–20, represent the averages of 77, 175, 36, 59, 32, 39, 50, 102, 121, 40, 59, 163, 26, 59, 79, 159, 211, 50, 46, and 15 particle images, respectively. Bar, 7 nm. C and D, Group averages of the core complex that showed one and two pores, respectively, were merged, yielding two main groups that were subjected to further alignment, classification, and averaging. The maps shown in C and D were calculated from 306 and 866 particles, respectively. Bar, 7 nm. E, Survey view of trypsin-treated core complex. Bar, 11 nm. F, Classification analysis of 777 trypsin-treated TOM core complex particles. The data set was split into 20 classes, as in B. Classes 1–10 represent the averages of 51, 59, 131, 93, 120, 88, 34, 40, 35 and 102 particle images, respectively. Class averages of <10 particle images are not shown. Bar, 7 nm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2169338&req=5

Figure 7: EM and projection map of the TOM core complex. A, Survey view of negatively stained TOM core complex. The image was filtered to the first zero of the electron microscope contrast transfer function. Bar, 11 nm. B, Classification analysis of 1,598 TOM core complex particles. Using MSA, the data set was split into 20 classes. Classes 1–20, represent the averages of 77, 175, 36, 59, 32, 39, 50, 102, 121, 40, 59, 163, 26, 59, 79, 159, 211, 50, 46, and 15 particle images, respectively. Bar, 7 nm. C and D, Group averages of the core complex that showed one and two pores, respectively, were merged, yielding two main groups that were subjected to further alignment, classification, and averaging. The maps shown in C and D were calculated from 306 and 866 particles, respectively. Bar, 7 nm. E, Survey view of trypsin-treated core complex. Bar, 11 nm. F, Classification analysis of 777 trypsin-treated TOM core complex particles. The data set was split into 20 classes, as in B. Classes 1–10 represent the averages of 51, 59, 131, 93, 120, 88, 34, 40, 35 and 102 particle images, respectively. Class averages of <10 particle images are not shown. Bar, 7 nm.

Mentions: Electron micrographs of negatively stained TOM core complex particles displayed predominantly two stain filled openings or pores, but particles representing a single ring were also present (Fig. 7 A). The length of the two pore particles was ∼12 nm, with a width of ∼7 nm. For further image processing, a total of 1,598 particle images were extracted and aligned with respect to translation and rotation via cross-correlation (Frank et al. 1981). Using MSA (Frank and van Heel 1982), 30 eigenimages were calculated and the data set was broken up into 20 classes using the 10 most significant eigenimages. The class averages contained predominantly two or one pores (Fig. 7 B). Preparations of the TOM holo complex contained roughly equal amounts of particles with either two or three rings (Künkele et al. 1998). None of the three ring structures were observed in the core complex preparations. Group averages that showed one and two pores, respectively, were merged, yielding two main groups. Classification analysis was then used to eliminate remaining core complexes with poorly defined structures. This analysis resulted in projection maps of two core complex classes (Fig. 7C and Fig. D).


The TOM core complex: the general protein import pore of the outer membrane of mitochondria.

Ahting U, Thun C, Hegerl R, Typke D, Nargang FE, Neupert W, Nussberger S - J. Cell Biol. (1999)

EM and projection map of the TOM core complex. A, Survey view of negatively stained TOM core complex. The image was filtered to the first zero of the electron microscope contrast transfer function. Bar, 11 nm. B, Classification analysis of 1,598 TOM core complex particles. Using MSA, the data set was split into 20 classes. Classes 1–20, represent the averages of 77, 175, 36, 59, 32, 39, 50, 102, 121, 40, 59, 163, 26, 59, 79, 159, 211, 50, 46, and 15 particle images, respectively. Bar, 7 nm. C and D, Group averages of the core complex that showed one and two pores, respectively, were merged, yielding two main groups that were subjected to further alignment, classification, and averaging. The maps shown in C and D were calculated from 306 and 866 particles, respectively. Bar, 7 nm. E, Survey view of trypsin-treated core complex. Bar, 11 nm. F, Classification analysis of 777 trypsin-treated TOM core complex particles. The data set was split into 20 classes, as in B. Classes 1–10 represent the averages of 51, 59, 131, 93, 120, 88, 34, 40, 35 and 102 particle images, respectively. Class averages of <10 particle images are not shown. Bar, 7 nm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169338&req=5

Figure 7: EM and projection map of the TOM core complex. A, Survey view of negatively stained TOM core complex. The image was filtered to the first zero of the electron microscope contrast transfer function. Bar, 11 nm. B, Classification analysis of 1,598 TOM core complex particles. Using MSA, the data set was split into 20 classes. Classes 1–20, represent the averages of 77, 175, 36, 59, 32, 39, 50, 102, 121, 40, 59, 163, 26, 59, 79, 159, 211, 50, 46, and 15 particle images, respectively. Bar, 7 nm. C and D, Group averages of the core complex that showed one and two pores, respectively, were merged, yielding two main groups that were subjected to further alignment, classification, and averaging. The maps shown in C and D were calculated from 306 and 866 particles, respectively. Bar, 7 nm. E, Survey view of trypsin-treated core complex. Bar, 11 nm. F, Classification analysis of 777 trypsin-treated TOM core complex particles. The data set was split into 20 classes, as in B. Classes 1–10 represent the averages of 51, 59, 131, 93, 120, 88, 34, 40, 35 and 102 particle images, respectively. Class averages of <10 particle images are not shown. Bar, 7 nm.
Mentions: Electron micrographs of negatively stained TOM core complex particles displayed predominantly two stain filled openings or pores, but particles representing a single ring were also present (Fig. 7 A). The length of the two pore particles was ∼12 nm, with a width of ∼7 nm. For further image processing, a total of 1,598 particle images were extracted and aligned with respect to translation and rotation via cross-correlation (Frank et al. 1981). Using MSA (Frank and van Heel 1982), 30 eigenimages were calculated and the data set was broken up into 20 classes using the 10 most significant eigenimages. The class averages contained predominantly two or one pores (Fig. 7 B). Preparations of the TOM holo complex contained roughly equal amounts of particles with either two or three rings (Künkele et al. 1998). None of the three ring structures were observed in the core complex preparations. Group averages that showed one and two pores, respectively, were merged, yielding two main groups. Classification analysis was then used to eliminate remaining core complexes with poorly defined structures. This analysis resulted in projection maps of two core complex classes (Fig. 7C and Fig. D).

Bottom Line: It forms a double ring structure that, in contrast to the holo complex, lacks the third density seen in the latter particles.Three-dimensional reconstruction by electron tomography exhibits two open pores traversing the complex with a diameter of approximately 2.1 nm and a height of approximately 7 nm.Tom40 is the key structural element of the TOM core complex.

View Article: PubMed Central - PubMed

Affiliation: Institut für Physiologische Chemie der Universität München, D-80336 München, Germany.

ABSTRACT
Translocation of nuclear-encoded preproteins across the outer membrane of mitochondria is mediated by the multicomponent transmembrane TOM complex. We have isolated the TOM core complex of Neurospora crassa by removing the receptors Tom70 and Tom20 from the isolated TOM holo complex by treatment with the detergent dodecyl maltoside. It consists of Tom40, Tom22, and the small Tom components, Tom6 and Tom7. This core complex was also purified directly from mitochondria after solubilization with dodecyl maltoside. The TOM core complex has the characteristics of the general insertion pore; it contains high-conductance channels and binds preprotein in a targeting sequence-dependent manner. It forms a double ring structure that, in contrast to the holo complex, lacks the third density seen in the latter particles. Three-dimensional reconstruction by electron tomography exhibits two open pores traversing the complex with a diameter of approximately 2.1 nm and a height of approximately 7 nm. Tom40 is the key structural element of the TOM core complex.

Show MeSH