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- Magnusdottir, Audur, et al.
(författare)
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The structure of the PP2A regulatory subunit B56gamma : The remaining piece of the PP2A jigsaw puzzle
- 2009
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 74:1, s. 212-221
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Tidskriftsartikel (refereegranskat)abstract
- The PP2A serine/threonine phosphatase regulates a plethora of cellular processes. In the cell the predominant form of the enzyme is a heterotrimer, formed by a core dimer composed of a catalytic and a scaffolding subunit, which assemble together with one of a range of different regulatory B subunits. Here, we present the first structure of a free non-complexed B subunit, B56. Comparison with the recent structures of a heterotrimeric complex and the core dimer reveals several significant conformational changes in the interface region between the B56 and the core dimer. These allow for an assembly scheme of the PP2A holoenzyme to be put forth where B56 first complexes with the scaffolding subunit and subsequently binds to the catalytic subunit and this induces the formation of a binding site for the invariant C-terminus of the catalytic subunit that locks in the complex as a last step of assembly.
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| 2. |
- Stenmark, Pål, 1976-
(författare)
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Structural and Functional Studies of Diiron Carboxylate Proteins
- 2005
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Iron is essential to all life; it is a vital component of many proteins in humans as well as plants and bacteria. Because of the ability of iron to activate oxygen, it is often found in proteins that interact with oxygen in some way. One of the protein families that utilize iron to harness the oxidative power of the oxygen molecule for different purposes is the diiron carboxylate protein family. These proteins have two iron ions bound in the active site that are coordinated by four carboxylic residues and two histidines. These enzymes catalyse reactions such as radical generation, ubiquinol oxidation, fatty acid desaturation, iron oxidation and many different hydroxylation reactions. One subgroup of this protein family that recently has been discovered contains the membrane-bound diiron carboxylate proteins. In this thesis we have studied three diiron carboxylate proteins. The first, alternative oxidase, is a membrane-bound ubiquinol oxidase present in the respiratory chain of plants, some yeasts and protozoa. We have obtained the first spectroscopical evidence for the presence of a diiron carboxylate site in alternative oxidase. We have also identified the first prokaryotic alternative oxidase and shown it to be expressed and functional. The second membrane-bound diiron carboxylate protein studied is Coq7; the inactivation of this protein has been reported to prolong the life span of the model organism Caenorhabditis elegans. We have shown that Coq7 catalyses a hydroxylation in the biosynthesis of ubiquinone and identified it as a diiron carboxylate protein. Finally we have solved the structure of the ribonucleotide reductase R2 subunit from Chlamydia trachomatis, a soluble diiron carboxylate protein. We have discovered that this is an unusual member of the R2 family because it challenges the generally accepted dogma of a conserved radical-harbouring tyrosine being a requirement for enzyme activity in this protein class. We have by EPR spectroscopy demonstrated that the radical in this protein is stored as a high-valent species at the diiron site instead of on a tyrosine. We also found that many organisms, including several human pathogens, have R2 proteins with the same sequence abnormalities and propose that these proteins constitute a new ribonucleotide reductase subclass, class Ic.
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| 3. |
- Stenmark, Pål, et al.
(författare)
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The Crystal Structure of the Bifunctional Deaminase/Reductase RibD of the Riboflavin Biosynthetic Pathway in Escherichia coli: Implications for the Reductive Mechanism
- 2007
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 373:1, s. 48-64
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Tidskriftsartikel (refereegranskat)abstract
- We have determined the crystal structure of the bi-functional deaminase/reductase enzyme from Escherichia coli (EcRibD) that catalyzes two consecutive reactions during riboflavin biosynthesis. The polypeptide chain of EcRibD is folded into two domains where the 3D structure of the N-terminal domain (1–145) is similar to cytosine deaminase and the C-terminal domain (146–367) is similar to dihydrofolate reductase. We showed that EcRibD is dimeric and compared our structure to tetrameric RibG, an ortholog from Bacillus subtilis (BsRibG). We have also determined the structure of EcRibD in two binary complexes with the oxidized cofactor (NADP+) and with the substrate analogue ribose-5-phosphate (RP5) and superposed these two in order to mimic the ternary complex. Based on this superposition we propose that the invariant Asp200 initiates the reductive reaction by abstracting a proton from the bound substrate and that the pro-R proton from C4 of the cofactor is transferred to C1 of the substrate. A highly flexible loop is found in the reductase active site (159–173) that appears to control cofactor and substrate binding to the reductase active site and was therefore compared to the corresponding Met20 loop of E. coli dihydrofolate reductase (EcDHFR). Lys152, identified by comparing substrate analogue (RP5) coordination in the reductase active site of EcRibD with the homologous reductase from Methanocaldococcus jannaschii (MjaRED), is invariant among bacterial RibD enzymes and could contribute to the various pathways taken during riboflavin biosynthesis in bacteria and yeast.
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