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The structure of Tim50(164-361) suggests the mechanism by which Tim50 receives mitochondrial presequences.

Li J, Sha B - Acta Crystallogr F Struct Biol Commun (2015)

Bottom Line: Compared with the previously determined Tim50(164-361) structure, significant conformational changes occur within the protruding β-hairpin of Tim50 and the nearby helix A2.These findings indicate that the IMS domain of Tim50 exhibits significant structural plasticity within the putative presequence-binding groove, which may play important roles in the function of Tim50 as a receptor protein in the TIM complex that interacts with the presequence and multiple other proteins.Therefore, the protruding β-hairpin of Tim50 may play critical roles in receiving the presequence and recruiting Tim23 for subsequent protein translocations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell, Developmental and Integrative Biology (CDIB), University of Alabama at Birmingham, Birmingham, AL 35294, USA.

ABSTRACT
Mitochondrial preproteins are transported through the translocase of the outer membrane (TOM) complex. Tim50 and Tim23 then transfer preproteins with N-terminal targeting presequences through the intermembrane space (IMS) across the inner membrane. The crystal structure of the IMS domain of Tim50 [Tim50(164-361)] has previously been determined to 1.83 Å resolution. Here, the crystal structure of Tim50(164-361) at 2.67 Å resolution that was crystallized using a different condition is reported. Compared with the previously determined Tim50(164-361) structure, significant conformational changes occur within the protruding β-hairpin of Tim50 and the nearby helix A2. These findings indicate that the IMS domain of Tim50 exhibits significant structural plasticity within the putative presequence-binding groove, which may play important roles in the function of Tim50 as a receptor protein in the TIM complex that interacts with the presequence and multiple other proteins. More interestingly, the crystal packing indicates that helix A1 from the neighboring monomer docks into the putative presequence-binding groove of Tim50(164-361), which may mimic the scenario of Tim50 and the presequence complex. Tim50 may recognize and bind the presequence helix by utilizing the inner side of the protruding β-hairpin through hydrophobic interactions. Therefore, the protruding β-hairpin of Tim50 may play critical roles in receiving the presequence and recruiting Tim23 for subsequent protein translocations.

No MeSH data available.


Related in: MedlinePlus

The putative presequence-binding grooves of Tim50_new molecules are occupied by secondary structures from neighboring molecules generated by crystal packing. (a) Helix A1 from the neighboring molecule (monomer A) is docked into the putative presequence-binding groove of monomer E of the Tim50_new structure primarily by hydrophobic interactions. Monomer E of Tim50_new belongs to group I and is shown in gold. Helix A1 from the neighboring molecule is shown in light blue and is labeled A1′. The secondary structures A2, B2 and B3 of monomer E are labeled. The residues Tyr223, Tyr227 and Gln230 from A1′, residues Trp207 and Trp213 from B2 and B3 and residues Asn240 and Tyr244 from A2 that are involved in the interactions are labeled. The hydrogen bond between Gln230 from A1 and Asn240 from A2 is indicated by a dotted line. (b) The N-terminal proline-rich loop from the neighboring molecule is inserted into the putative presequence-binding groove of monomer A of the Tim50_new structure by hydrophobic interactions. Monomer A of the Tim50_new structure belongs to group II and is shown in cyan. The N-­terminal proline-rich loop from the neighboring molecule is shown in green. The residues Pro187 and Tyr188 from the N-terminal proline-rich loop, residues Trp207 and Trp213 from B2 and B3 and residue Tyr244 from A2 that are involved in the interactions are labeled.
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fig3: The putative presequence-binding grooves of Tim50_new molecules are occupied by secondary structures from neighboring molecules generated by crystal packing. (a) Helix A1 from the neighboring molecule (monomer A) is docked into the putative presequence-binding groove of monomer E of the Tim50_new structure primarily by hydrophobic interactions. Monomer E of Tim50_new belongs to group I and is shown in gold. Helix A1 from the neighboring molecule is shown in light blue and is labeled A1′. The secondary structures A2, B2 and B3 of monomer E are labeled. The residues Tyr223, Tyr227 and Gln230 from A1′, residues Trp207 and Trp213 from B2 and B3 and residues Asn240 and Tyr244 from A2 that are involved in the interactions are labeled. The hydrogen bond between Gln230 from A1 and Asn240 from A2 is indicated by a dotted line. (b) The N-terminal proline-rich loop from the neighboring molecule is inserted into the putative presequence-binding groove of monomer A of the Tim50_new structure by hydrophobic interactions. Monomer A of the Tim50_new structure belongs to group II and is shown in cyan. The N-­terminal proline-rich loop from the neighboring molecule is shown in green. The residues Pro187 and Tyr188 from the N-terminal proline-rich loop, residues Trp207 and Trp213 from B2 and B3 and residue Tyr244 from A2 that are involved in the interactions are labeled.

Mentions: It is interesting to note that the protruding β-hairpin and helix A2 exhibit different conformations even in structures from the same crystallization conditions. To further understand the mechanism by which the group I and group II members of Tim50_new adopt different conformations, we examined the neighboring molecules of the group I and group II members generated by crystal packing. We reasoned that the neighbouring environment of Tim50(164–361) may play an important role in the conformation that Tim50(164–361) adopts. Surprisingly, for members of both group I and group II the putative presequence-binding grooves are occupied by secondary structures from the neighboring monomers. For group I members helix A1 from the neighboring monomer is docked into the putative presequence-binding groove, and for group II members the N-terminal proline-rich loop from the neighboring monomer is inserted into the putative pre­sequence-binding groove (Fig. 3 ▸). In sharp contrast, when we examined the crystal packing in the Tim50_old structure, the putative presequence-binding groove is virtually empty (data not shown). Therefore, it is likely that Tim50_old represents the presequence-free conformation of the IMS domain of Tim50 and the Tim50_new corresponds to the presequence-binding conformation of the IMS domain of Tim50. The structural differences between the members of group I and group II within one asymmetric unit of the Tim50_new structure suggests that the putative presequence-binding groove of the IMS domain of Tim50 may adopt different conformations to accommodate various presequences.


The structure of Tim50(164-361) suggests the mechanism by which Tim50 receives mitochondrial presequences.

Li J, Sha B - Acta Crystallogr F Struct Biol Commun (2015)

The putative presequence-binding grooves of Tim50_new molecules are occupied by secondary structures from neighboring molecules generated by crystal packing. (a) Helix A1 from the neighboring molecule (monomer A) is docked into the putative presequence-binding groove of monomer E of the Tim50_new structure primarily by hydrophobic interactions. Monomer E of Tim50_new belongs to group I and is shown in gold. Helix A1 from the neighboring molecule is shown in light blue and is labeled A1′. The secondary structures A2, B2 and B3 of monomer E are labeled. The residues Tyr223, Tyr227 and Gln230 from A1′, residues Trp207 and Trp213 from B2 and B3 and residues Asn240 and Tyr244 from A2 that are involved in the interactions are labeled. The hydrogen bond between Gln230 from A1 and Asn240 from A2 is indicated by a dotted line. (b) The N-terminal proline-rich loop from the neighboring molecule is inserted into the putative presequence-binding groove of monomer A of the Tim50_new structure by hydrophobic interactions. Monomer A of the Tim50_new structure belongs to group II and is shown in cyan. The N-­terminal proline-rich loop from the neighboring molecule is shown in green. The residues Pro187 and Tyr188 from the N-terminal proline-rich loop, residues Trp207 and Trp213 from B2 and B3 and residue Tyr244 from A2 that are involved in the interactions are labeled.
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fig3: The putative presequence-binding grooves of Tim50_new molecules are occupied by secondary structures from neighboring molecules generated by crystal packing. (a) Helix A1 from the neighboring molecule (monomer A) is docked into the putative presequence-binding groove of monomer E of the Tim50_new structure primarily by hydrophobic interactions. Monomer E of Tim50_new belongs to group I and is shown in gold. Helix A1 from the neighboring molecule is shown in light blue and is labeled A1′. The secondary structures A2, B2 and B3 of monomer E are labeled. The residues Tyr223, Tyr227 and Gln230 from A1′, residues Trp207 and Trp213 from B2 and B3 and residues Asn240 and Tyr244 from A2 that are involved in the interactions are labeled. The hydrogen bond between Gln230 from A1 and Asn240 from A2 is indicated by a dotted line. (b) The N-terminal proline-rich loop from the neighboring molecule is inserted into the putative presequence-binding groove of monomer A of the Tim50_new structure by hydrophobic interactions. Monomer A of the Tim50_new structure belongs to group II and is shown in cyan. The N-­terminal proline-rich loop from the neighboring molecule is shown in green. The residues Pro187 and Tyr188 from the N-terminal proline-rich loop, residues Trp207 and Trp213 from B2 and B3 and residue Tyr244 from A2 that are involved in the interactions are labeled.
Mentions: It is interesting to note that the protruding β-hairpin and helix A2 exhibit different conformations even in structures from the same crystallization conditions. To further understand the mechanism by which the group I and group II members of Tim50_new adopt different conformations, we examined the neighboring molecules of the group I and group II members generated by crystal packing. We reasoned that the neighbouring environment of Tim50(164–361) may play an important role in the conformation that Tim50(164–361) adopts. Surprisingly, for members of both group I and group II the putative presequence-binding grooves are occupied by secondary structures from the neighboring monomers. For group I members helix A1 from the neighboring monomer is docked into the putative presequence-binding groove, and for group II members the N-terminal proline-rich loop from the neighboring monomer is inserted into the putative pre­sequence-binding groove (Fig. 3 ▸). In sharp contrast, when we examined the crystal packing in the Tim50_old structure, the putative presequence-binding groove is virtually empty (data not shown). Therefore, it is likely that Tim50_old represents the presequence-free conformation of the IMS domain of Tim50 and the Tim50_new corresponds to the presequence-binding conformation of the IMS domain of Tim50. The structural differences between the members of group I and group II within one asymmetric unit of the Tim50_new structure suggests that the putative presequence-binding groove of the IMS domain of Tim50 may adopt different conformations to accommodate various presequences.

Bottom Line: Compared with the previously determined Tim50(164-361) structure, significant conformational changes occur within the protruding β-hairpin of Tim50 and the nearby helix A2.These findings indicate that the IMS domain of Tim50 exhibits significant structural plasticity within the putative presequence-binding groove, which may play important roles in the function of Tim50 as a receptor protein in the TIM complex that interacts with the presequence and multiple other proteins.Therefore, the protruding β-hairpin of Tim50 may play critical roles in receiving the presequence and recruiting Tim23 for subsequent protein translocations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cell, Developmental and Integrative Biology (CDIB), University of Alabama at Birmingham, Birmingham, AL 35294, USA.

ABSTRACT
Mitochondrial preproteins are transported through the translocase of the outer membrane (TOM) complex. Tim50 and Tim23 then transfer preproteins with N-terminal targeting presequences through the intermembrane space (IMS) across the inner membrane. The crystal structure of the IMS domain of Tim50 [Tim50(164-361)] has previously been determined to 1.83 Å resolution. Here, the crystal structure of Tim50(164-361) at 2.67 Å resolution that was crystallized using a different condition is reported. Compared with the previously determined Tim50(164-361) structure, significant conformational changes occur within the protruding β-hairpin of Tim50 and the nearby helix A2. These findings indicate that the IMS domain of Tim50 exhibits significant structural plasticity within the putative presequence-binding groove, which may play important roles in the function of Tim50 as a receptor protein in the TIM complex that interacts with the presequence and multiple other proteins. More interestingly, the crystal packing indicates that helix A1 from the neighboring monomer docks into the putative presequence-binding groove of Tim50(164-361), which may mimic the scenario of Tim50 and the presequence complex. Tim50 may recognize and bind the presequence helix by utilizing the inner side of the protruding β-hairpin through hydrophobic interactions. Therefore, the protruding β-hairpin of Tim50 may play critical roles in receiving the presequence and recruiting Tim23 for subsequent protein translocations.

No MeSH data available.


Related in: MedlinePlus