| 1. |
- Van Lun, Michiel, et al.
(author)
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CO2 and O-2 Distribution in Rubisco Suggests the Small Subunit Functions as a CO2 Reservoir
- 2014
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In: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 0002-7863 .- 1520-5126. ; 136:8, s. 3165-3171
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Journal article (peer-reviewed)abstract
- Protein gas interactions are important in biology. The enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyzes two competing reactions involving CO2 and O-2 as substrates. Carboxylation of the common substrate ribulose-1,5-bisphosphate leads to photosynthetic carbon assimilation, while the oxygenation reaction competes with carboxylation and reduces photosynthetic productivity. The migration of the two gases in and around Rubisco was investigated using molecular dynamics simulations. The results indicate that at equal concentrations of the gases, Rubisco binds CO2 stronger than it does O-2. Amino acids with small hydrophobic side chains are the most proficient in attracting CO2, indicating a significant contribution of the hydrophobic effect in the interaction. On average, residues in the small subunit bind approximately twice as much CO2 as do residues in the large subunit. We did not detect any cavities that would provide a route to the active site for the gases. Instead, CO2 appears to be guided toward the active site through a CO2 binding region around the active site opening that extends to the closest neighboring small subunits. Taken together, these results suggest the small subunit may function as a "reservoir" for CO2 storage.
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| 2. |
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| 3. |
- Van Lun, Michiel, et al.
(author)
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Subunit Interface Dynamics in Hexadecameric Rubisco
- 2011
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In: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 411:5, s. 1083-1098
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Journal article (peer-reviewed)abstract
- Ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase (Rubisco) plays an important role in the global carbon cycle as a hub for biomass. Rubisco catalyzes not only the carboxylation of RuBP with carbon dioxide but also a competing oxygenation reaction of RuBP with a negative impact on photosynthetic yield. The functional active site is built from two large (L) subunits that form a dimer. The octameric core of four L(2) dimers is held at each end by a cluster of four small (S) subunits, forming a hexadecamer. Each large subunit contacts more than one S subunit. These interactions exploit the dynamic flexibility of Rubisco, which we address in this study. Here, we describe seven different types of interfaces of hexadecameric Rubisco. We have analyzed these interfaces with respect to the size of the interface area and the number of polar interactions, including salt bridges and hydrogen bonds in a variety of Rubisco enzymes from different organisms and different kingdoms of life, including the Rubisco-like proteins. We have also performed molecular dynamics simulations of Rubisco from Chlamydomonas reinhardtii and mutants thereof. From our computational analyses, we propose structural checkpoints of the S subunit to ensure the functionality and/or assembly of the Rubisco holoenzyme. These checkpoints appear to fine-tune the dynamics of the enzyme in a way that could influence enzyme performance.
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| 4. |
- Valegård, Karin, et al.
(author)
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Structural and functional analyses of Rubisco from arctic diatom species reveal unusual posttranslational modifications
- 2018
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In: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 293:34, s. 13033-13043
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Journal article (peer-reviewed)abstract
- The catalytic performance of the major CO2-assimilating enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), restricts photosynthetic productivity. Natural diversity in the catalytic properties of Rubisco indicates possibilities for improvement. Oceanic phytoplankton contain some of the most efficient Rubisco enzymes, and diatoms in particular are responsible for a significant proportion of total marine primary production as well as being a major source of CO2 sequestration in polar cold waters. Until now, the biochemical properties and three-dimensional structures of Rubisco from diatoms were unknown. Here, diatoms from arctic waters were collected, cultivated, and analyzed for their CO2-fixing capability. We characterized the kinetic properties of five and determined the crystal structures of four Rubiscos selected for their high CO2-fixing efficiency. The DNA sequences of the rbcL, and rbcS genes of the selected diatoms were similar, reflecting their close phylogenetic relationship. The V-max and K-m for the oxygenase and carboxylase activities at 25 degrees C and the specificity factors (S-c/o) at 15, 25, and 35 degrees C were determined. The S-c/o values were high, approaching those of mono- and dicot plants, thus exhibiting good selectivity for CO(2 )relative to O-2. Structurally, diatom Rubiscos belong to form I C/D, containing small subunits characterized by a short beta A-beta B loop and a C-terminal extension that forms a beta-hairpin structure (beta E-beta F loop). Of note, the diatom Rubiscos featured a number of posttranslational modifications of the large subunit, including 4-hydroxyproline, beta-hydroxyleucine, hydroxylated and nitrosylated cysteine, mono- and dihydroxylated lysine, and trimethylated lysine. Our studies suggest adaptation toward achieving efficient CO2 fixation in arctic diatom Rubiscos.
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| 5. |
- Van Lun, Michiel
(author)
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Structural dynamics of ribulose-1,5-bisphosphate carboxylase/oxygenase
- 2013
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Doctoral thesis (other academic/artistic)abstract
- Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) assimilates carbon dioxide (CO2) from air into biomass. Due to its slow turnover, the reaction is a rate-limiting step in photosynthetic carbon fixation. The carboxylation reaction catalyzed by Rubisco is subject to inhibition by oxygen (O2) in a competing, non-productive reaction that reduces the efficiency of the enzyme by up to 50%. This makes Rubisco a target for engineering to increase crop yield. The specificity of Rubisco for CO2 over O2 is a measure how well the enzyme is able to suppress the unwanted oxygenation reaction and varies between organisms. The specificity of Rubisco from several marine algae surpasses that of crop plants. Diatoms with high CO2 specificity from the arctic waters around Svalbard have been cultured, the Rubisco protein has been isolated and characterised, and the crystal structure has been determined. The holoenzyme structure is similar to the structure of Rubisco from plants, but the fold of the small subunits differs and has a shorter βA-βB loop and carboxy- terminal extension that extends into the solvent channel, that appears to provide extra stability to the holoenzyme. The holoenzyme is a hexadecamer consisting of 8 large, catalytic, and 8 small subunits (L8S8) with a mass of 500 kD. The dynamics of the interaction between the subunits in this large protein will likely influence catalysis and CO2/O2 specificity. In order to examine the interface communication between subunits, molecular dynamics simulations have been performed on Rubisco enzymes from different organisms and with different holoenzyme structures, showing that the number of contacts and the size of the interaction area differ significantly in the different complexes examined. Single-residue mutations that affect specificity in Rubisco from the unicellular green alga Chlamydomonas reinhardtii also influence the protein dynamics and interactions across the subunit interfaces. The migration of the gaseous substrates, CO2 and O2 in and around Rubisco, was investigated using molecular dynamics simulations. The results indicate that at equal concentrations of the gas, Rubisco has a preference for binding CO2 over O2. Amino acids with small hydrophobic side chains are the most proficient in attracting CO2, indicating a significant contribution of the hydrophobic effect in the interaction. On average, residues in the small subunit bind approximately twice as much CO2 as do residues in the large subunit, suggesting the small subunit may function as a reservoir for CO2 storage.
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