Structural insight into the SAM-mediated assembly of the mitochondrial TOM core complex

Abstract

Barrels that build barrels Cells produce specialized machinery to produce functional β-barrel proteins that are involved in cross-membrane transport and membrane protein biogenesis in eukaryotic organelles and some bacteria. Wang et al . studied one of these systems that is itself a β-barrel, the mitochondrial sorting and assembly machinery (SAM), in complex with one of its client proteins, translocase of the outer membrane (TOM). TOM subunits can be added one at a time while the protein is bound to SAM. Compared with the full TOM core complex, the final TOM core subunit clashes with SAM and suggests that release of TOM may be driven by its association. —MAF

Targeting and Insertion of Membrane Proteins in Mitochondria

“…This has the effect of limiting the amount of Mdm10 which is able to bind to the SAM complex thus inhibiting TOM complex assembly. Cryo-EM experiments have recently generated high resolution structures of the SAM complex from M. thermophila and S. cerevisiae in complex with various substrates providing more evidence to support the mechanisms of outer membrane protein insertion discussed above (Diederichs et al, 2020;Takeda et al, 2021;Wang et al, 2021). The structural data clearly show a lateral opening in Sam50 between the β-signal in strand 16 and strand 1.…”

“…The outer membrane therefore represents a significant barrier for these proteins which they must cross in order to be correctly assembled into both the inner and outer mitochondrial membranes. The entry gate that controls this import process is known as the TOM complex and is composed of a β-barrel pore forming protein (Tom40), a number of accessory/ scaffolding proteins (Tom5, Tom6 and Tom7) and two receptor proteins (Tom20 and Tom70) that recognise mitochondrial targeting signals within protein sequences (Bolliger et al, 1995;Dietmeier et al, 1997;Rapaport and Neupert, 1999;Model et al, 2001;Gabriel et al, 2003;Mokranjac and Neupert, 2015;Pfanner et al, 2019;Wang et al, 2021). Additionally, the TOM complex contains a protein called Tom22, which appears to act as both a receptor and a scaffolding protein helping to control the number of pore-forming subunits associated to each fully assembled TOM complex as well as facilitating protein import (Lithgow et al, 1994;Bolliger et al, 1995;Moczko et al, 1997;Yano et al, 2000).…”

“…(Kiebler et al, 1990), however it is only with recent advances in techniques such as cryo-EM that the assembly pathway of the TOM complex has become clearer. The assembly of the TOM complex is separated into three distinct stages known as assembly I, assembly II and TOM core assembly (Wang et al, 2021) (Figure 2A). Assembly I involves the integration of the Tom40 β-barrel protein into the outer membrane from the intermembrane space (IMS) side of the outer membrane via the sorting and assembly machinery (SAM) complex (this process will be discussed in detail later) (Wiedemann et al, 2003).…”

“…Tom5 and Tom6 are subsequently inserted and assembled with Tom40 while it is still associated with the SAM complex (Dietmeier et al, 1997;Model et al, 2001;Becker et al, 2010;Thornton et al, 2010). The subsequent addition of Tom7 leads to dissociation of the growing TOM complex from SAM and constitutes assembly II (Yamano et al, 2010;Becker et al, 2011b;Wang et al, 2021). The addition of Tom22 to the complex leads to the assembly of the TOM core complex containing multiple fully assembled TOM complexes with Tom22 acting as a scaffold holding them together (Araiso et al, 2019;Tucker and Park, 2019;Wang et al, 2020b;Wang et al, 2021).…”

“…Sam35 is thought to be peripherally associated with the SAM complex facing the cytosol which is counterintuitive to a mechanistic understanding given that β-barrel proteins are inserted from the IMS side of the outer membrane. There is some evidence that Sam35 is actually embedded within the outer membrane through close interactions with Sam50 (Kutik et al, 2008), possibly within the pore of Sam50, although recent structural data do not support this hypothesis (Takeda et al, 2021;Wang et al, 2021).…”