Mitochondrial protein import: Identification of processing peptidase and of PEP, a processing enhancing protein

Hawlitschek, Gerhard; Schneider, Hans‐Jürgen; Schmidt, Bernd; Tropschug, Maximilian; Hartl, F. Ulrich; Neupert, Walter

doi:10.1016/0092-8674(88)90096-7

Cited by 376 publications

(191 citation statements)

References 62 publications

(61 reference statements)

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“…ATP is needed independently of the requirement for a membrane potential [31]. Recently, the processing peptidase of the mitochondrial matrix and a processing enhancing protein ('PEP'), which is largely associated with the inner membrane, were identified [32]. A major advance in studying the distinct steps of protein import came from the reversible accumulation of precursor proteins at defined stages of their import pathway ('translocation arrest').…”

Section: Reviewmentioning

confidence: 99%

Import of proteins into mitochondria: a multi‐step process

Pfanner

Hartl

Neupert

1988

European Journal of Biochemistry

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148

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Translocation of precursor proteins from the cytosol into mitochondria is a multi-step process. The generation of translocation intermediates, i.e. the reversible accumulation of precursors at distinct stages of their import pathway into mitochondria ('translocation arrest'), has allowed the experimental characterization of distinct functional steps of protein import. These steps include: ATP-dependent unfolding of precursors; specific recognition of precursors by distinct receptors on the mitochondrial surface; interaction of precursors with a general insertion protein ('GIP') in the outer mitochondrial membrane; membrane-potential-dependent translocation into the inner membrane at contact sites between both membranes; proteolytic processing of precursors; and intramitochondrial sorting of precursors via the matrix space ('conservative sorting'). The functional characteristics unveiled by studying mitochondrial protein import appear to be of general interest for investigations on intracellular protein sorting.Over 90% of mitochondrial proteins are nuclear-coded and are synthesized as precursor proteins on cytosolic polysomes (for recent reviews, see [l -31). The precursors are then translocated to their functional destination in one of the four mitochondrial compartments (outer membrane, intermembrane space, inner membrane, and matrix). More than ten years ago, it was demonstrated that mitochondrial protein import could occur post-translationally in vivo and in vitro, thereby proving the mechanistic independence of translation and translocation, and that protein import required energy [4-71. Since then, several important characteristics of mitochondrial protein uptake have been unravelled which we would like to mention in a short historical overview. These include the demonstration of amino-terminal peptide extensions ('presequences') on the precursor proteins [S, 91 which are proteolytically processed in the mitochondrial matrix [lo]. On the other hand, several precursors have been shown to by synthesized without cleavable peptide extensions [ l l , 121. Protein translocation into the inner membrane requires the membrane potential ( A Y) across the inner mitochondrial membrane [13 -151. Several precursors are proteolytically processed in two steps, the second processing activity residing in the interrnembrane space [16-IS]. The primary sequences of precursor proteins have been determined, demonstrating that the presequences are positively charged [19, 201. Precursors imported in vitro have been shown to be assembled into multi-subunit protein complexes [21,22]. Amino-terminal precursor sequences, either the presequences or amino-terminal portions in the mature part of non-cleavable precursors, have

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Section: Reviewmentioning

confidence: 99%

Import of proteins into mitochondria: a multi‐step process

Pfanner

Hartl

Neupert

1988

European Journal of Biochemistry

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148

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“…Most proteins destined for import into the mitochondrial matrix bear targeting information at their N termini. These matrix targeting sequences (MTSs) are commonly 20 -60 amino acids in length and are cleaved upon import into mitochondria by the mitochondrial processing peptidase (MPP) (4,5). N-terminal presequences contain several positively charged amino acids, are notably devoid of negative charges, and contain periodic hydroxylated amino acid residues.…”

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confidence: 99%

The DNA Helicase, Hmi1p, Is Transported into Mitochondria by a C-terminal Cleavable Targeting Signal

Lee

Sedman

Neupert

et al. 1999

Journal of Biological Chemistry

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104

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We have identified a novel mitochondrial targeting signal in the precursor of the DNA helicase Hmi1p of Saccharomyces cerevisiae that is located at the C terminus of the protein. Similar to classical N-terminal presequences, this C-terminal targeting signal consists of a stretch of positively charged amino acids that has the potential to form an amphipathic ␣-helix. Deletion of the C-terminal 36 amino acids of helicase resulted in loss of import into mitochondria, while deletion of the N-terminal 40 amino acids had no effect. When C-terminal regions of the helicase were placed at the C terminus of a passenger protein, dihydrofolate reductase, the resulting fusion proteins were directed into the mitochondrial matrix, and the C-terminal region of helicase became proteolytically processed. Import of helicase occurs in a C-to N-terminal direction; it requires a membrane potential and the TIM17-23 translocase together with mitochondrial Hsp70. Helicase is the only mitochondrial matrix protein identified thus far with a cleavable targeting signal at its C terminus.

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“…A major breakthrough was the demonstration of an ATP-dependent protease with a similarity to E. coli Lon (or La) in the matrix of mitochondria from rat liver (Desautels andGoldberg 1982a, 1985), counterparts of which were later found in bovine adrenal cortex (Watabe and Kimura 1985a,b) and yeast (Kutejová et al 1993). In addition, non-ATP-dependent metalloproteases involved in the processing of precursor proteins were identified in mitochondria from rat liver (Conboy et al 1982;Miura et al 1982), bovine adrenal cortex (Kumamoto et al 1986), yeast (McAda andDouglas 1982;Böhni et al 1983), and N. crassa (Hawlitchek et al 1988). Only in the last few years have genes that encode these and other (putative) mitochondrial proteases been isolated from S. cerevisiae and other organisms ( Table 2).…”

Section: Mitochondrial Proteasesmentioning

confidence: 99%