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Iwakura, Masahiro; Nakamura, Tsutomu; Yamane, Chiori; Maki, Kosuke

doi:10.1038/76811

Cited by 121 publications

(101 citation statements)

References 27 publications

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“…Mitochondrial precursor proteins were constructed by fusing varying lengths of the N-terminal portion of precytochrome b 2 to the N termini of B. amyloliquefaciens ribonuclease barnase (5), the E. coli chemotaxis response regulator CheY (11), or circular permutants (CPs) of E. coli dihydrofolate reductase (DHFR) (12) in pGEM-3Zf(ϩ) vectors. The CheY mutational analysis was conducted in F14N͞ V54T double mutant as pseudo wild type into which the mutations V11T, V33T, A42G, V83T, A98G, and A103G were introduced (11).…”

Section: Methodsmentioning

confidence: 99%

“…In these experiments, the original N and C termini of DHFR were connected by a short linker, new N termini were created at Pro-25 (CP P25) or Lys-38 (CP K38), and mitochondrial targeting sequences were fused to the proteins at the new N termini (26). The DHFR mutants have similar structures and stabilities and differ primarily in their topology (12). The amino acids immediately adjacent to the N terminus of CP P25 DHFR form an ␣-helix at the surface of the protein, as does the N terminus of barnase.…”

Section: The Local Structure Adjacent To the Targeting Sequence Affectsmentioning

confidence: 99%

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Effect of protein structure on mitochondrial import

Wilcox

Choy

Bustamante

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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Most proteins that are to be imported into the mitochondrial matrix are synthesized as precursors, each composed of an Nterminal targeting sequence followed by a mature domain. Precursors are recognized through their targeting sequences by receptors at the mitochondrial surface and are then threaded through import channels into the matrix. Both the targeting sequence and the mature domain contribute to the efficiency with which proteins are imported into mitochondria. Precursors must be in an unfolded conformation during translocation. Mitochondria can unfold some proteins by changing their unfolding pathways. The effectiveness of this unfolding mechanism depends on the local structure of the mature domain adjacent to the targeting sequence. This local structure determines the extent to which the unfolding pathway can be changed and, therefore, the unfolding rate increased. Atomic force microscopy studies find that the local structures of proteins near their N and C termini also influence their resistance to mechanical unfolding. Thus, protein unfolding during import resembles mechanical unfolding, and the specificity of import is determined by the resistance of the mature domain to unfolding as well as by the properties of the targeting sequence.protein unfolding ͉ mitochondria ͉ protein import ͉ protein sorting ͉ atomic force microscopy

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Section: Methodsmentioning

confidence: 99%

Section: The Local Structure Adjacent To the Targeting Sequence Affectsmentioning

confidence: 99%

Effect of protein structure on mitochondrial import

Wilcox

Choy

Bustamante

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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“…Several proteins have been permuted circularly and retain a similar native structure, including T4 lysozyme (16) (20), and dihydrofolate reductase (21). Two of these, ␣-spectrin SH3 (22) and CI2 (23), fold with two-state kinetics, and both maintain a two-state folding mechanism after permutation.…”

Section: Folding Algorithms Based On the Topology Of The Native Statementioning

confidence: 99%

Experimental evaluation of topological parameters determining protein-folding rates

Miller

Fischer²,

Marqusee³

2002

Proc. Natl. Acad. Sci. U.S.A.

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Recent work suggests that structural topology plays a key role in determining protein-folding rates and pathways. The refolding rates of small proteins that fold without intermediates are found to correlate with simple structural parameters such as relative contact order, long-range order, or the fraction of short-range contacts. To test and evaluate the role of structural topology experimentally, a set of circular permutants of the ribosomal protein S6 from Thermus thermophilus was analyzed. Despite a wide range of relative contact order, the permuted proteins all fold with similar rates. These results suggest that alternative topological parameters may better describe the role of topology in proteinfolding rates.T he amino acid sequence of a protein and its chemical environment determine its native structure (1), but how this structure is determined from sequence remains one of the major unsolved problems in biology. The process of protein folding cannot be accomplished by random search through all possible conformations, because the number of structures available to an unfolded protein is too large; therefore there must be a biased or directional search in order for a protein to reach its native state. Small Two-State Proteins Show a Wide Range in Folding RatesThe simplest models for studying the protein-folding process are those that refold without intermediates (2). To date there are more than 20 examples of such simple systems. These proteins all fold cooperatively in a mono-exponential fashion from the denatured state, but the rates at which they fold span 6 orders of magnitude. What causes this remarkable variety of refolding rates? Folding Algorithms Based on the Topology of the Native State Have Predictive ValueRecently, several folding algorithms based on native structural information have been used to predict folding rates and nucleation sites (3-6). These efforts suggest that the topology of the final structure is an important determinant in the mechanism of protein folding.If native topology plays a major role in protein folding, is there a simple structural parameter that will capture this feature and explain the large range of observed folding rates? Recently, several simple parameters defining topological features of the native state have been shown to correlate well with proteinrefolding rates. The first and most commonly used parameter is relative contact order (RCO; ref. 7). Remarkably, RCO, which represents the normalized average sequence separation between contacting residues, was observed to correlate extremely well with the folding rates of a small set of two-state proteins. The lower the RCO, the faster the protein folds. Although it is possible to alter folding rates significantly through point mutations that do not change RCO, and there are proteins with similar RCOs and different folding rates (8), in general this correlation has improved as more two-state proteins have been characterized (9) and can explain the difference in folding rates among a family of structurally homologous pro...

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“…for E. coli DHFR and ⌬G unfolding ϭ 4.4 kcal mol Ϫ1 for mouse DHFR (61,62)). Thus, as the proteases encountered the DHFR domain, some of the time they would succeed in unfolding the DHFR domain and degrade the substrate completely, and some of the time they would fail to unfold the DHFR domain, and the remainder of the substrate would dissociate to yield a partially degraded substrate (Scheme 1).…”

mentioning

confidence: 99%

ATP-dependent Proteases Differ Substantially in Their Ability to Unfold Globular Proteins

Koodathingal

Jaffe

Kraut

et al. 2009

Journal of Biological Chemistry

114

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ATP-dependent proteases control the concentrations of hundreds of regulatory proteins and remove damaged or misfolded proteins from cells. They select their substrates primarily by recognizing sequence motifs or covalent modifications. Once a substrate is bound to the protease, it has to be unfolded and translocated into the proteolytic chamber to be degraded. Some proteases appear to be promiscuous, degrading substrates with poorly defined targeting signals, which suggests that selectivity may be controlled at additional levels. Here we compare the abilities of representatives from all classes of ATP-dependent proteases to unfold a model substrate protein and find that the unfolding abilities range over more than 2 orders of magnitude. We propose that these differences in unfolding abilities contribute to the fates of substrate proteins and may act as a further layer of selectivity during protein destruction.Energy-dependent proteolysis is responsible for more than 90% of the protein turnover inside the cell (1). This process both removes misfolded and aggregated proteins as part of the response of the cell to stress and controls the concentrations of regulatory proteins (2, 3). In prokaryotes and eukaryotic organelles, energy-dependent proteases fall into five classes as follows: ClpAP, ClpXP, Lon, HslUV (also referred to as ClpYQ), and HflB (also referred to as FtsH). In Archaea, analogous functions are performed by the archaebacterial proteasome, consisting of the proteasome-activating nucleotidase (PAN), 3 working with the 20 S proteasome (4); in the cytoplasm and nucleus of eukaryotes, these same functions are performed by the 26 S proteasome (5). These different proteases show little sequence conservation outside the ATP-binding domains, but they share their overall architecture. They all form oligomeric, barrel-shaped complexes composed of one or more rings with the active sites of proteolysis sequestered inside a central degradation chamber (6). Access channels to these sites are narrow, and proteins have to be unfolded to gain entry (6). Regulatory particles belonging to the AAA family of molecular chaperones assemble on either end of the proteolytic chamber and recognize substrates destined for degradation. After recognition, the regulatory particles translocate the substrate through a central channel to the proteolytic chamber and in doing so unravel folded domains within the substrate. Translocation and unfolding are driven by ATP hydrolysis by the regulatory particles, with conformational changes in the protease transmitted to the substrate by conserved residues in the loops lining the channel (7-10).Protein degradation by AAA proteases is tightly regulated. Most proteins are targeted to ClpAP, ClpXP, HslUV, Lon, HflB, and PAN by sequence motifs in their primary structure (11)(12)(13)(14)(15)(16)(17). Sometimes adaptor proteins recognize and bind sequence elements in substrates and deliver them to the protease, and other times the protease recognizes sequence elements directly (18,19). In contrast,...

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Cited by 121 publications

References 27 publications

Effect of protein structure on mitochondrial import

Effect of protein structure on mitochondrial import

Experimental evaluation of topological parameters determining protein-folding rates

ATP-dependent Proteases Differ Substantially in Their Ability to Unfold Globular Proteins

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