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Mitochondrial protein translocase (MPT) (Chacinska et al., 2005; Mokranjac et al., 2005; Bihlmaier et al., 2007). The crystal structure of the intermembrane space domain of yeast Tim50 has been solved to 1.83 Å resolution (Qian et al., 2011). A protruding beta-hairpin of Tim50 is crucial for interaction with Tim23, providing a molecular basis for the cooperation of Tim50 and Tim23 in preprotein translocation to the protein-conducting channel of the mitochondrial inner membrane (Qian et al., 2011).  TIM23-mediates insertion of transmembrane α-helices into the mitochondrial inner membrane (Botelho et al., 2011). The TIM23 channel undergoes structural changes in response to the energized state of the membrane, the pmf (Malhotra et al. 2013).  TMS1 in TIM23 is required for homodimerization while it and TMS2 are involved in pre-protein binding in the channel (Pareek et al. 2013).  The Tom40 outer membrane channel may be a 19 β-stranded barrel, possibly homologous to the VDAC porins (TC# 1.B.8) (Lackey et al. 2014).  Tim23 and Tim17 interact with each other as well as Tim44 and Pam17, respectively.  These last two proteins may serve regulatory functions (Ting et al. 2014).  Tom20, 22, 40 and 70 recognize presequences in various mitochondrially targetted proteins (Melin et al. 2015; Melin et al. 2014).  In the 4 TMS TIM17 protein, mutations in TMSs1 and 2 impair the interaction of Tim17 with Tim23, whereas mutations in TM3 compromise binding of the import motor (Demishtein-Zohary et al. 2017); further, residues in the matrix-facing region of Tim17 involved in binding of the import motor were identified. TIM22, forms an intramolecular disulfide bond in yeast and humans.  If not oxidized, they do not properly integrate into the membrane complex, and the lack of Tim17 oxidation disrupts the TIM23 translocase complex (Wrobel et al. 2016). Mgr2 (TC# 1.A.111.1.3) and Pam18 are involved in precursormembrane protein quality control (Schendzielorz et al. 2018). Tom7 and OMA1 play reciprocal roles during mitochondrial import and activation of the PTEN-induced kinase 1, PINK1, in humans (Sekine et al. 2019).  Organellar beta-barrel proteins are unique as most of them do not contain typical targeting information in the form of an N-terminal cleavable targeting signal. Instead, targeting and surface recognition of mitochondrial beta-barrel proteins in yeast, humans and plants depends on the hydrophobicity of the last beta-hairpin of the beta-barrel. Klinger et al. 2019 demonstrated that hydrophobicity is not sufficient for the discrimination of targeting to chloroplasts or mitochondria. Using atVDAC1 (TC# 1.B.8.1.15) and psOEP24 (1.B.28.1.1) they showed that the presence of a hydrophilic amino acid at the C-terminus of the penultimate beta-strand is required for mitochondrial targeting. A mutation of the chloroplast beta-barrel protein psOEP24 which mimics such a profile is efficiently targeted to mitochondria (Klinger et al. 2019). The high-resolution cryo-EM structures of the core TOM complex from Saccharomyces cerevisiae in dimeric and tetrameric forms have  been determined (Tucker and Park 2019). Dimeric TOM consists of two copies each of five proteins arranged in two-fold symmetry: pore-forming beta-barrel protein Tom40 and four auxiliary alpha-helical transmembrane proteins. The pore of each Tom40 has an overall negatively charged inner surface due to multiple functionally important acidic patches. The tetrameric complex is a dimer of dimeric TOM, which may be capable of forming higher-order oligomers. Negatively charged residues in the N-terminus of Tim17 are critical for the preprotein-induced gating of the TIM23 translocase, possibly by recognizing the positive charges in the leader sequence of the substrate proteins (Meier et al. 2005). Tom22, Tom40 and Tom7 have been observed in primorial eukaryotes, arguing that these subunits were core to the complex early in evolution (Maćasev et al. 2004). The assembly of the Tim22 complex have been studied () Kumar et al. 2020. The intermembrane space (IMS) and TMS4 regions of Tim22 are required for interactions with membrane-embedded subunits, including Tim54, Tim18, and Sdh3, and thereby maintain the functional architecture of the TIM22 translocase. The TMS1 and TMS2 regions of Tim22 are important for association with Tim18, whereas TMS3 is exclusively required for the interaction with Sdh3 (Kumar et al. 2020). TIM8.13 and TIM9.10 are chaparone proteins that form complexes, Tim23/TIM8.13 and Tim23/TIM9.10 complexes. TIM8.13 uses transient salt bridges to interact with the hydrophilic part of its client, but its interactions to the transmembrane part are weaker than in TIM9.10. Consequently, TIM9.10 outcompetes TIM8.13 in binding hydrophobic clients, while TIM8.13 is tuned to a few clients with both hydrophilic and hydrophobic parts (Sučec et al. 2020). The TOM complex is a multisubunit membrane protein complex consisting of a beta-barrel protein Tom40 and six alpha-helical transmembrane (TM) proteins, receptor subunits Tom20, Tom22, and Tom70, and regulatory subunits Tom5, Tom6, and Tom7 (Araiso et al. 2020). CryoEM structures showed a symmetric dimer containing five different subunits including Tom22. Different translocation paths within the Tom40 import channel seem to be present for different classes of translocating precursor proteins (Araiso et al. 2020). The Mgr2 subunit of the TIM23 complex regulates membrane insertion of marginal stop-transfer signals in the mitochondrial inner membrane (Lee et al. 2020). The architecture of the human TIM22 complex has been examined by chemical crosslinking (Valpadashi et al. 2021).


MPT of Saccharomyces cerevisiae Outer Membrane Translocase
Tom5 (channel regulator)
Tom6 (channel regulator)
Tom7 (channel regulator)
Tom20 (receptor)
Tom22 (receptor)
Tom40 (channel) (involved in sorting)
Tom70 (receptor) Inner Membrane Translocase #1 (Tim23 complex)
Tim17 (channel protein)
Tim23 (channel protein)
Tim44 (ATP-dependent motor)
Tim14 (J protein; PAM18; Ylr008c) (motor component)
Tim16 (Pam16) (motor component)
Tim21 (may regulate protein import by binding to both the translocase of the outer membrane (TOM) and presequence-associated motor (PAM) complexes)
Mge1 (motor component)
Mdj2 (J protein that can replace Tim14) Intermembrane Chaperone for the Tim23 Complex
Tim50 Inner Membrane Translocase #2 (Tim22 complex)
Tim22 (channel protein)
Tim54 (channel protein)
Tim18 (channel protein) Intermembrane Chaperones for the Tim22 Complex
Tim8 (targeting protein)
Tim9 (targeting protein)
Tim10 (targeting protein)
Tim13 (targeting protein) Intermembrane (IM) Proteins involved in uniport and assembly of other intermembrane proteins, Mia40 and Erv1 (Chacinska et al., 2004; Grumbt et al., 2007))
Mia40 (403aas)
Erv1 (P27882)

Hydrogenosome multicomponent protein import complex.

Import complex of Trichomonas vaginalis
Small TIM9-10A, 74 aas, A2FKF0
Small TIM9-10B, 74 aas, A2DZ11
Tim17A/B protein, 160 aas, A2DDM1
Tim17C, 159 aas, A2FHN8
Tim17D, 157 aas, 4 TMSs, A2DXD3
Tim17-like, 170 aas, A2DS29
Tim44, 326 aas, A2F7V1
Pam16, 111 aas, A2FE18
Pam18, 117 aas, A2DFA6

The mitochondrial Tim17/Tim 22/Tim23 protein import system. This complex mediates the translocation of transit peptide-containing proteins across the mitochondrial inner membrane and links the inner and outer membranes.  It plays a role in germination (Wang et al. 2014).

Protein import complex of Arabidopsis thaliana (Mouse-ear cress)
Tim17 (Tim17-1) of 243 aas and 3 TMSs (Q9SP35)
Tim22 (Tim22-1; MEE67) of 173 aas and 3 TMSs (A2RVP7)
Tim23 (Tim23-1; Tha1) of 187 aas and 3 TMSs (Q9LNQ1)

The chloroplast Tic17-2/Tic22-2/Tic23-2 complex. It brings proteins into the chloroplast (Teng et al. 2006; Kasmati et al. 2011).

The Tic17/Tic22/Tic23 complex of Arabidopsis thaliana (Mouse-ear cress)
Tic17-2, 243 aas and 3 or 4 TMSs, Q9SP35
Tic22-2 (HP20), 210 aas and 4 TMSs, Q94EH2
Tic23-2, 188 aas and 3 or 4 TMSs, Q38820