| 1. |
- Charavgi, Maria-Despoina, et al.
(författare)
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The structure of a novel glucuronoyl esterase from Myceliophthora thermophila gives new insights into its role as a potential biocatalyst
- 2013
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Ingår i: Acta Crystallographica Section D. - 0907-4449 .- 1399-0047. ; 69:1, s. 63-73
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Tidskriftsartikel (refereegranskat)abstract
- The increasing demand for the development of efficient biocatalysts is a consequence of their broad industrial applications. Typical difficulties that are encountered during their exploitation in a variety of processes are interconnected with factors such as temperature, pH, product inhibitors etc. To eliminate these, research has been directed towards the identification of new enzymes that would comply with the required standards. To this end, the recently discovered glucuronoyl esterases (GEs) are an enigmatic family within the carbohydrate esterase (CE) family. Structures of the thermophilic StGE2 esterase from Myceliophthora thermophila (synonym Sporotrichum thermophile), a member of the CE15 family, and its S213A mutant were determined at 1.55 and 1.9 Å resolution, respectively. The first crystal structure of the S213A mutant in complex with a substrate analogue, methyl 4-O-methyl-[beta]-D-glucopyranuronate, was determined at 2.35 Å resolution. All of the three-dimensional protein structures have an [alpha]/[beta]-hydrolase fold with a three-layer [alpha][beta][alpha]-sandwich architecture and a Rossmann topology and comprise one molecule per asymmetric unit. These are the first crystal structures of a thermophilic GE both in an unliganded form and bound to a substrate analogue, thus unravelling the organization of the catalytic triad residues and their neighbours lining the active site. The knowledge derived offers novel insights into the key structural elements that drive the hydrolysis of glucuronic acid esters.
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| 2. |
- Chrysina, Maria, et al.
(författare)
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Five-coordinate Mn-IV intermediate in the activation of nature's water splitting cofactor
- 2019
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 116:34, s. 16841-16846
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Tidskriftsartikel (refereegranskat)abstract
- Nature's water splitting cofactor passes through a series of catalytic intermediates (S-0-S-4) before O-O bond formation and O-2 release. In the second last transition (S-2 to S-3) cofactor oxidation is coupled to water molecule binding to Mn1. It is this activated, water-enriched all Mn-IV form of the cofactor that goes on to form the O-O bond, after the next light-induced oxidation to S-4. How cofactor activation proceeds remains an open question. Here, we report a so far not described intermediate (S-3') in which cofactor oxidation has occurred without water insertion. This intermediate can be trapped in a significant fraction of centers (> 50%) in (i) chemical-modified cofactors in which Ca2+ is exchanged with Sr2+; the Mn4O5Sr cofactor remains active, but the S-2-S-3 and S-3-S-0 transitions are slower than for the Mn4O5Ca cofactor; and (ii) upon addition of 3% vol/vol methanol; methanol is thought to act as a substrate water analog. The S-3' electron paramagnetic resonance (EPR) signal is significantly broader than the untreated S-3 signal (2.5 T vs. 1.5 T), indicating the cofactor still contains a 5-coordinate Mn ion, as seen in the preceding S-2 state. Magnetic double resonance data extend these findings revealing the electronic connectivity of the S-3' cofactor is similar to the high spin form of the preceding S-2 state, which contains a cuboidal Mn3O4Ca unit tethered to an external, 5-coordinate Mn ion (Mn-4). These results demonstrate that cofactor oxidation regulates water molecule insertion via binding to Mn-4. The interaction of ammonia with the cofactor is also discussed.
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| 3. |
- Dimarogona, Maria, et al.
(författare)
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The crystal structure of a Fusarium oxysporum feruloyl esterase that belongs to the tannase family
- 2020
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Ingår i: FEBS Letters. - : John Wiley & Sons. - 0014-5793 .- 1873-3468. ; 594:11, s. 1738-1749
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Tidskriftsartikel (refereegranskat)abstract
- Feruloyl esterases are enzymes of industrial interest that catalyse the hydrolysis of the ester bond between hydroxycinnamic acids such as ferulic acid and sugars present in the plant cell wall. Although there are several structures of biochemically characterized feruloyl esterases available, the structural determinants of their substrate specificity are not yet fully understood. Here, we present the crystal structure of a feruloyl esterase from Fusarium oxysporum (FoFaeC) at 2.3 Å resolution. Similar to the two other tannase‐like feruloyl esterases, FoFaeC features a large lid domain covering the active site with potential regulatory role and a disulphide bond that brings together the serine and histidine of the catalytic triad. Differences are mainly observed in the metal coordination site and the substrate binding pocket.
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