The Experts below are selected from a list of 10143 Experts worldwide ranked by ideXlab platform
Xiaojiang S Chen - One of the best experts on this subject based on the ideXlab platform.
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mechanisms of conformational change for a replicative hexameric helicase of sv40 large tumor antigen
Cell, 2004Co-Authors: Xiaojiang S Chen, Dahai Gai, Rui Zhao, Carla V FinkielsteinAbstract:Abstract The large tumor antigen (LTag) of simian virus 40, an AAA + Protein, is a hexameric helicase essential for viral DNA replication in eukaryotic cells. LTag functions as an efficient molecular machine powered by ATP binding and hydrolysis for origin DNA melting and replication fork unwinding. To understand how ATP binding and hydrolysis are coupled to conformational changes, we have determined high-resolution structures (∼1.9 A) of LTag hexamers in distinct nucleotide binding states. The structural differences of LTag in various nucleotide states detail the molecular mechanisms of conformational changes triggered by ATP binding/hydrolysis and reveal a potential mechanism of concerted nucleotide binding and hydrolysis. During these conformational changes, the angles and orientations between domains of a monomer alter, creating an "iris"-like motion in the hexamer. Additionally, six unique β hairpins on the channel surface move longitudinally along the central channel, possibly serving as a motor for pulling DNA into the LTag double hexamer for unwinding.
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mechanisms of conformational change for a replicative hexameric helicase of sv40 large tumor antigen
Cell, 2004Co-Authors: Dahai Gai, Xiaojiang S Chen, Rui Zhao, Carla V FinkielsteinAbstract:The large tumor antigen (LTag) of simian virus 40, an AAA(+) Protein, is a hexameric helicase essential for viral DNA replication in eukaryotic cells. LTag functions as an efficient molecular machine powered by ATP binding and hydrolysis for origin DNA melting and replication fork unwinding. To understand how ATP binding and hydrolysis are coupled to conformational changes, we have determined high-resolution structures ( approximately 1.9 A) of LTag hexamers in distinct nucleotide binding states. The structural differences of LTag in various nucleotide states detail the molecular mechanisms of conformational changes triggered by ATP binding/hydrolysis and reveal a potential mechanism of concerted nucleotide binding and hydrolysis. During these conformational changes, the angles and orientations between domains of a monomer alter, creating an "iris"-like motion in the hexamer. Additionally, six unique beta hairpins on the channel surface move longitudinally along the central channel, possibly serving as a motor for pulling DNA into the LTag double hexamer for unwinding.
Feng Wang - One of the best experts on this subject based on the ideXlab platform.
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structural dynamics of the meca clpc complex a type ii AAA Protein unfolding machine
Journal of Biological Chemistry, 2013Co-Authors: Ningning Li, Yutao Qi, Yanji Xu, Feng WangAbstract:Abstract The MecA-ClpC complex is a bacterial type II AAA+ molecular machine responsible for regulated unfolding of substrates, such as transcription factors ComK and ComS, and targeting them to ClpP for degradation. The six subunits of the MecA-ClpC complex form a closed barrel-like structure, featured with three stacked rings and a hollow passage, where substrates are threaded and translocated through successive pores. Although the general concepts of how polypeptides are unfolded and translocated by internal pore loops of AAA+ Proteins have long been conceived, the detailed mechanistic model remains elusive. With cryoelectron microscopy, we captured four different structures of the MecA-ClpC complexes. These complexes differ in the nucleotide binding states of the two AAA+ rings and therefore might presumably reflect distinctive, representative snapshots from a dynamic unfolding cycle of this hexameric complex. Structural analysis reveals that nucleotide binding and hydrolysis modulate the hexameric complex in a number of ways, including the opening of the N-terminal ring, the axial and radial positions of pore loops, the compactness of the C-terminal ring, as well as the relative rotation between the two nucleotide-binding domain rings. More importantly, our structural and biochemical data indicate there is an active allosteric communication between the two AAA+ rings and suggest that concerted actions of the two AAA+ rings are required for the efficiency of the substrate unfolding and translocation. These findings provide important mechanistic insights into the dynamic cycle of the MecA-ClpC unfoldase and especially lay a foundation toward the complete understanding of the structural dynamics of the general type II AAA+ hexamers.
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structural dynamics of the meca clpc complex a type ii AAA Protein unfolding machine
Journal of Biological Chemistry, 2013Co-Authors: Jing Liu, Feng Wang, Ziqing Mei, Yigong Shi, Jianlin Lei, Ning GaoAbstract:The MecA-ClpC complex is a bacterial type II AAA+ molecular machine responsible for regulated unfolding of substrates, such as transcription factors ComK and ComS, and targeting them to ClpP for degradation. The six subunits of the MecA-ClpC complex form a closed barrel-like structure, featured with three stacked rings and a hollow passage, where substrates are threaded and translocated through successive pores. Although the general concepts of how polypeptides are unfolded and translocated by internal pore loops of AAA+ Proteins have long been conceived, the detailed mechanistic model remains elusive. With cryoelectron microscopy, we captured four different structures of the MecA-ClpC complexes. These complexes differ in the nucleotide binding states of the two AAA+ rings and therefore might presumably reflect distinctive, representative snapshots from a dynamic unfolding cycle of this hexameric complex. Structural analysis reveals that nucleotide binding and hydrolysis modulate the hexameric complex in a number of ways, including the opening of the N-terminal ring, the axial and radial positions of pore loops, the compactness of the C-terminal ring, as well as the relative rotation between the two nucleotide-binding domain rings. More importantly, our structural and biochemical data indicate there is an active allosteric communication between the two AAA+ rings and suggest that concerted actions of the two AAA+ rings are required for the efficiency of the substrate unfolding and translocation. These findings provide important mechanistic insights into the dynamic cycle of the MecA-ClpC unfoldase and especially lay a foundation toward the complete understanding of the structural dynamics of the general type II AAA+ hexamers. Background: The structure of the type II AAA+ hexameric molecular machine is highly dynamic. Results: Nucleotide binding and hydrolysis induce twisted movements of pore loops and relative rotation between two AAA+ rings. Conclusion: An allosteric communication between two AAA+ rings is present in type II AAA+ hexamer. Significance: This work provides a comprehensive understanding of the structural dynamics of type II AAA+ hexamer.
H Schuler - One of the best experts on this subject based on the ideXlab platform.
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crystal structure of the atpase domain of the human AAA Protein paraplegin spg7
PLOS ONE, 2009Co-Authors: T Karlberg, Susanne Van Den Berg, Martin Hammarstrom, J Sagemark, Ida Johansson, L Holmbergschiavone, H SchulerAbstract:Paraplegin is an m-AAA protease of the mitochondrial inner membrane that is linked to hereditary spastic paraplegias. The gene encodes an FtsH-homology protease domain in tandem with an AAA+ homology ATPase domain. The Protein is believed to form a hexamer that uses ATPase-driven conformational changes in its AAA-domain to deliver substrate peptides to its protease domain. We present the crystal structure of the AAA-domain of human paraplegin bound to ADP at 2.2 A. This enables assignment of the roles of specific side chains within the catalytic cycle, and provides the structural basis for understanding the mechanism of disease mutations.
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crystal structure of the atpase domain of the human AAA Protein paraplegin spg7
PLOS ONE, 2009Co-Authors: T Karlberg, Martin Hammarstrom, J Sagemark, Ida Johansson, L Holmbergschiavone, Susanne Van Den Berg, H SchulerAbstract:Paraplegin is an m-AAA protease of the mitochondrial inner membrane that is linked to hereditary spastic paraplegias. The gene encodes an FtsH-homology protease domain in tandem with an AAA+ homology ATPase domain. The Protein is believed to form a hexamer that uses ATPase-driven conformational changes in its AAA-domain to deliver substrate peptides to its protease domain. We present the crystal structure of the AAA-domain of human paraplegin bound to ADP at 2.2 A. This enables assignment of the roles of specific side chains within the catalytic cycle, and provides the structural basis for understanding the mechanism of disease mutations. Enhanced version This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.
Manajit Hayerhartl - One of the best experts on this subject based on the ideXlab platform.
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rubisco activases AAA chaperones adapted to enzyme repair
Frontiers in Molecular Biosciences, 2017Co-Authors: J Y Bhat, Ulrich F Hartl, Gabriel Thieulinpardo, Manajit HayerhartlAbstract:Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), the key enzyme of the Calvin-Benson-Bassham cycle of photosynthesis, requires conformational repair by Rubisco activase for efficient function. Rubisco mediates the fixation of atmospheric CO2 by catalyzing the carboxylation of the five-carbon sugar ribulose-1,5-bisphosphate (RuBP). It is a remarkably inefficient enzyme, and efforts to increase crop yields by bioengineering Rubisco remain unsuccessful. This is due in part to the complex cellular machinery required for Rubisco biogenesis and metabolic maintenance. To function, Rubisco must undergo an activation process that involves carboxylation of an active site lysine by a non-substrate CO2 molecule and binding of a Mg2+ ion. Premature binding of the substrate RuBP results in an inactive enzyme. Moreover, Rubisco can also be inhibited by a range of sugar phosphates, some of which are ‘misfire’ products of its multistep catalytic reaction. The release of the inhibitory sugar molecule is mediated by the AAA+ Protein Rubisco activase (Rca), which couples hydrolysis of ATP to the structural remodeling of Rubisco. Rca enzymes are found in the vast majority of photosynthetic organisms, from bacteria to higher plants. They share a canonical AAA+ domain architecture and form six-membered ring complexes but are diverse in sequence and mechanism, suggesting their convergent evolution. In this review, we discuss recent advances in understanding the structure and function of this important group of client-specific AAA+ Proteins.
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degradation of potent rubisco inhibitor by selective sugar phosphatase
Nature plants, 2015Co-Authors: Andreas Bracher, Ulrich F Hartl, A Sharma, A Starlingwindhof, Manajit HayerhartlAbstract:Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyses the conversion of atmospheric carbon dioxide into organic compounds in photosynthetic organisms. Alongside carboxylating the five-carbon sugar ribulose-1,5-bisphosphate (RuBP)(1-3), Rubisco produces a small amount of xylulose-1,5-bisphosphate (XuBP), a potent inhibitor of Rubisco(4). The AAA+ Protein Rubisco activase removes XuBP from the active site of Rubisco in an ATP-dependent process(5,6). However, free XuBP rapidly rebinds to Rubisco, perpetuating its inhibitory effect. Here, we combine biochemical and structural analyses to show that the CbbY Protein of the photosynthetic bacterium Rhodobacter sphaeroides and Arabidopsis thaliana is a highly selective XuBP phosphatase. We also show that CbbY converts XuBP to the non-inhibitory compound xylulose-5-phosphate, which is recycled back to RuBP. We solve the crystal structures of CbbY from R. sphaeroides and A. thaliana, and through mutational analysis show that the cap domain of the Protein confers the selectivity for XuBP over RuBP. Finally, in vitro experiments with CbbY from R. sphaeroides reveal that CbbY cooperates with Rubisco activase to prevent a detrimental build-up of XuBP at the Rubisco active site. We suggest that CbbY, which is conserved in algae and plants, is an important component of the cellular machinery that has evolved to deal with the shortcomings of the ancient enzyme Rubisco.
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structure of green type rubisco activase from tobacco
Nature Structural & Molecular Biology, 2011Co-Authors: Mathias Stotz, Oliver Muellercajar, Andreas Bracher, Petra Wendler, Susanne Ciniawsky, F U Hartl, Manajit HayerhartlAbstract:Rubisco, the enzyme that catalyzes the fixation of atmospheric CO(2) in photosynthesis, is subject to inactivation by inhibitory sugar phosphates. Here we report the 2.95-A crystal structure of Nicotiana tabacum Rubisco activase (Rca), the enzyme that facilitates the removal of these inhibitors. Rca from tobacco has a classical AAA(+)-Protein domain architecture. Although Rca populates a range of oligomeric states when in solution, it forms a helical arrangement with six subunits per turn when in the crystal. However, negative-stain electron microscopy of the active mutant R294V suggests that Rca functions as a hexamer. The residues determining species specificity for Rubisco are located in a helical insertion of the C-terminal domain and probably function in conjunction with the N-domain in Rubisco recognition. Loop segments exposed toward the central pore of the hexamer are required for the ATP-dependent remodeling of Rubisco, resulting in the release of inhibitory sugar.
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structure and function of the AAA Protein cbbx a red type rubisco activase
Nature, 2011Co-Authors: Oliver Muellercajar, Andreas Bracher, Ulrich F Hartl, Mathias Stotz, Petra Wendler, Manajit HayerhartlAbstract:Ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyses the fixation of atmospheric CO(2) in photosynthesis, but tends to form inactive complexes with its substrate ribulose 1,5-bisphosphate (RuBP). In plants, Rubisco is reactivated by the AAA(+) (ATPases associated with various cellular activities) Protein Rubisco activase (Rca), but no such Protein is known for the Rubisco of red algae. Here we identify the Protein CbbX as an activase of red-type Rubisco. The 3.0-A crystal structure of unassembled CbbX from Rhodobacter sphaeroides revealed an AAA(+) Protein architecture. Electron microscopy and biochemical analysis showed that ATP and RuBP must bind to convert CbbX into functionally active, hexameric rings. The CbbX ATPase is strongly stimulated by RuBP and Rubisco. Mutational analysis suggests that CbbX functions by transiently pulling the carboxy-terminal peptide of the Rubisco large subunit into the hexamer pore, resulting in the release of the inhibitory RuBP. Understanding Rubisco activation may facilitate efforts to improve CO(2) uptake and biomass production by photosynthetic organisms.
Oliver Muellercajar - One of the best experts on this subject based on the ideXlab platform.
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insights into the mechanism and regulation of the cbbqo type rubisco activase a moxr AAA atpase
Proceedings of the National Academy of Sciences of the United States of America, 2020Co-Authors: Yichin Candace Tsai, Lynette Liew, Di Liu, Shashi Bhushan, Yonggui Gao, Oliver MuellercajarAbstract:The vast majority of biological carbon dioxide fixation relies on the function of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco). In most cases the enzyme exhibits a tendency to become inhibited by its substrate RuBP and other sugar phosphates. The inhibition is counteracted by diverse molecular chaperones known as Rubisco activases (Rcas). In some chemoautotrophic bacteria, the CbbQO-type Rca Q2O2 repairs inhibited active sites of hexameric form II Rubisco. The 2.2-A crystal structure of the MoxR AAA+ Protein CbbQ2 from Acidithiobacillus ferrooxidans reveals the helix 2 insert (H2I) that is critical for Rca function and forms the axial pore of the CbbQ hexamer. Negative-stain electron microscopy shows that the essential CbbO adaptor Protein binds to the conserved, concave side of the CbbQ2 hexamer. Site-directed mutagenesis supports a model in which adenosine 5'-triphosphate (ATP)-powered movements of the H2I are transmitted to CbbO via the concave residue L85. The basal ATPase activity of Q2O2 Rca is repressed but strongly stimulated by inhibited Rubisco. The characterization of multiple variants where this repression is released indicates that binding of inhibited Rubisco to the C-terminal CbbO VWA domain initiates a signal toward the CbbQ active site that is propagated via elements that include the CbbQ α4-β4 loop, pore loop 1, and the presensor 1-β hairpin (PS1-βH). Detailed mechanistic insights into the enzyme repair chaperones of the highly diverse CO2 fixation machinery of Proteobacteria will facilitate their successful implementation in synthetic biology ventures.
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probing the rice rubisco rubisco activase interaction via subunit heterooligomerization
Proceedings of the National Academy of Sciences of the United States of America, 2019Co-Authors: Devendra Shivhare, Yichin Candace Tsai, Oliver MuellercajarAbstract:During photosynthesis the AAA+ Protein and essential molecular chaperone Rubisco activase (Rca) constantly remodels inhibited active sites of the CO2-fixing enzyme Rubisco (ribulose 1,5-bisphosphate carboxylase/oxygenase) to release tightly bound sugar phosphates. Higher plant Rca is a crop improvement target, but its mechanism remains poorly understood. Here we used structure-guided mutagenesis to probe the Rubisco-interacting surface of rice Rca. Mutations in Ser-23, Lys-148, and Arg-321 uncoupled adenosine triphosphatase and Rca activity, implicating them in the Rubisco interaction. Mutant doping experiments were used to evaluate a suite of known Rubisco-interacting residues for relative importance in the context of the functional hexamer. Hexamers containing some subunits that lack the Rubisco-interacting N-terminal domain displayed a ∼2-fold increase in Rca function. Overall Rubisco-interacting residues located toward the rim of the hexamer were found to be less critical to Rca function than those positioned toward the axial pore. Rca is a key regulator of the rate-limiting CO2-fixing reactions of photosynthesis. A detailed functional understanding will assist the ongoing endeavors to enhance crop CO2 assimilation rate, growth, and yield.
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mechanism of enzyme repair by the AAA chaperone rubisco activase
Molecular Cell, 2017Co-Authors: J Y Bhat, Oliver Muellercajar, Andreas Bracher, Ulrich F Hartl, Gabriel Thieulinpardo, Susanne Ciniawsky, Goran Milicic, Andrew Maxwell, John R Engen, Petra WendlerAbstract:How AAA+ chaperones conformationally remodel specific target Proteins in an ATP-dependent manner is not well understood. Here, we investigated the mechanism of the AAA+ Protein Rubisco activase (Rca) in metabolic repair of the photosynthetic enzyme Rubisco, a complex of eight large (RbcL) and eight small (RbcS) subunits containing eight catalytic sites. Rubisco is prone to inhibition by tight-binding sugar phosphates, whose removal is catalyzed by Rca. We engineered a stable Rca hexamer ring and analyzed its functional interaction with Rubisco. Hydrogen/deuterium exchange and chemical crosslinking showed that Rca structurally destabilizes elements of the Rubisco active site with remarkable selectivity. Cryo-electron microscopy revealed that Rca docks onto Rubisco over one active site at a time, positioning the C-terminal strand of RbcL, which stabilizes the catalytic center, for access to the Rca hexamer pore. The pulling force of Rca is fine-tuned to avoid global destabilization and allow for precise enzyme repair.
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structure of green type rubisco activase from tobacco
Nature Structural & Molecular Biology, 2011Co-Authors: Mathias Stotz, Oliver Muellercajar, Andreas Bracher, Petra Wendler, Susanne Ciniawsky, F U Hartl, Manajit HayerhartlAbstract:Rubisco, the enzyme that catalyzes the fixation of atmospheric CO(2) in photosynthesis, is subject to inactivation by inhibitory sugar phosphates. Here we report the 2.95-A crystal structure of Nicotiana tabacum Rubisco activase (Rca), the enzyme that facilitates the removal of these inhibitors. Rca from tobacco has a classical AAA(+)-Protein domain architecture. Although Rca populates a range of oligomeric states when in solution, it forms a helical arrangement with six subunits per turn when in the crystal. However, negative-stain electron microscopy of the active mutant R294V suggests that Rca functions as a hexamer. The residues determining species specificity for Rubisco are located in a helical insertion of the C-terminal domain and probably function in conjunction with the N-domain in Rubisco recognition. Loop segments exposed toward the central pore of the hexamer are required for the ATP-dependent remodeling of Rubisco, resulting in the release of inhibitory sugar.
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structure and function of the AAA Protein cbbx a red type rubisco activase
Nature, 2011Co-Authors: Oliver Muellercajar, Andreas Bracher, Ulrich F Hartl, Mathias Stotz, Petra Wendler, Manajit HayerhartlAbstract:Ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyses the fixation of atmospheric CO(2) in photosynthesis, but tends to form inactive complexes with its substrate ribulose 1,5-bisphosphate (RuBP). In plants, Rubisco is reactivated by the AAA(+) (ATPases associated with various cellular activities) Protein Rubisco activase (Rca), but no such Protein is known for the Rubisco of red algae. Here we identify the Protein CbbX as an activase of red-type Rubisco. The 3.0-A crystal structure of unassembled CbbX from Rhodobacter sphaeroides revealed an AAA(+) Protein architecture. Electron microscopy and biochemical analysis showed that ATP and RuBP must bind to convert CbbX into functionally active, hexameric rings. The CbbX ATPase is strongly stimulated by RuBP and Rubisco. Mutational analysis suggests that CbbX functions by transiently pulling the carboxy-terminal peptide of the Rubisco large subunit into the hexamer pore, resulting in the release of the inhibitory RuBP. Understanding Rubisco activation may facilitate efforts to improve CO(2) uptake and biomass production by photosynthetic organisms.
