Glutathione transferases immobilized on nanoporous alumina: Flow system kinetics, screening, and stability

Kjellander, Marcus; Mazari, Aslam M. A.; Boman, Mats; Mannervik, Bengt; Johansson, Gunnar

doi:10.1016/j.ab.2013.10.004

Cited by 13 publications

(7 citation statements)

References 21 publications

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“…Despite the modest differences in substrate selectivities reported for these isoforms on the basis of a limited range of substrates (Kjellander et al, 2014), the structures of DmGSTE6 and DmGSTE7 are nearly invariant and they are also very similar to the few other epsilon-class GSTs for which structures are available. Although it is expected that the GSHbinding sites would be structurally conserved in other GST classes, the substrate-binding sites (H-sites) might be expected to vary more than is observed here between these isoforms.…”

Section: Discussionmentioning

confidence: 95%

“…The genes for DmGSTE6 and DmGSTE7 were cloned into a pET vector as described elsewhere (Kjellander et al, 2014), The proteins (with hexahistidine tags at their N-termini) were expressed in Escherichia coli strain BL21(DE3) and purified by affinity chromatography. Briefly, 6 Â 500 ml E .coli cultures in Terrific Broth supplemented with 50 mg ml À1 kanamycin were incubated at 37 C and 200 rev min À1 .…”

Section: Protein Expression and Biophysical Characterizationmentioning

confidence: 99%

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Comparison of epsilon- and delta-class glutathioneS-transferases: the crystal structures of the glutathioneS-transferases DmGSTE6 and DmGSTE7 fromDrosophila melanogaster

Scian

Trong

Mazari

et al. 2015

Acta Crystallogr D Biol Crystallogr

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Cytosolic glutathione transferases (GSTs) comprise a large family of enzymes with canonical structures that diverge functionally and structurally among mammals, invertebrates and plants. Whereas mammalian GSTs have been characterized extensively with regard to their structure and function, invertebrate GSTs remain relatively unstudied. The invertebrate GSTs do, however, represent potentially important drug targets for infectious diseases and agricultural applications. In addition, it is essential to fully understand the structure and function of invertebrate GSTs, which play important roles in basic biological processes. Invertebrates harbor delta- and epsilon-class GSTs, which are not found in other organisms.Drosophila melanogasterGSTs (DmGSTs) are likely to contribute to detoxication or antioxidative stress during development, but they have not been fully characterized. Here, the structures of two epsilon-class GSTs fromDrosophila, DmGSTE6 and DmGSTE7, are reported at 2.1 and 1.5 Å resolution, respectively, and are compared with other GSTs to identify structural features that might correlate with their biological functions. The structures of DmGSTE6 and DmGSTE7 are remarkably similar; the structures do not reveal obvious sources of the minor functional differences that have been observed. The main structural difference between the epsilon- and delta-class GSTs is the longer helix (A8) at the C-termini of the epsilon-class enzymes.

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Section: Discussionmentioning

confidence: 95%

Section: Protein Expression and Biophysical Characterizationmentioning

confidence: 99%

Comparison of epsilon- and delta-class glutathioneS-transferases: the crystal structures of the glutathioneS-transferases DmGSTE6 and DmGSTE7 fromDrosophila melanogaster

Scian

Trong

Mazari

et al. 2015

Acta Crystallogr D Biol Crystallogr

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“…Candida antarctica lipase B Hydrolysis of p-nitrophenyl palmitate Alumina [9] Rat hepatic microsomal CYP2E1 Para-nitrophenol hydroxylation Alumina [15] Alliinase Conversion of alliin in alliicin Alumina [16] Glucose-6-phosphate dehydrogenase Oxidation of Glucose-6 phosphate (G6P) Alumina [17] Silica [18] Laccase Oxidation of 2,2 1 -azino-bis-(3 ethylbenzothiazoline-6-sulfonic acid) Titania [19] Urease Urea hydroxylation Alumina [20] Glutathione transferase Glutathione conjugation to 1-chloro-2,4-dinitrobenzene Alumina [21] One of the most studied biocatalysts is lipase, which is active in different reactions such as hydrolysis, esterification, and transesterification, with high enantioselective properties [22]. Lipase is an interfacial molecule with phase transfer catalytic properties, meaning it performs to the best of its abilities in an oil/water interface [23].…”

Section: Immobilized Proteinmentioning

confidence: 99%

Use of a Ceramic Membrane to Improve the Performance of Two-Separate-Phase Biocatalytic Membrane Reactor

Ranieri

Mazzei

et al. 2016

Molecules

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Biocatalytic membrane reactors (BMR) combining reaction and separation within the same unit have many advantages over conventional reactor designs. Ceramic membranes are an attractive alternative to polymeric membranes in membrane biotechnology due to their high chemical, thermal and mechanical resistance. Another important use is their potential application in a biphasic membrane system, where support solvent resistance is highly needed. In this work, the preparation of asymmetric ceramic hollow fibre membranes and their use in a two-separate-phase biocatalytic membrane reactor will be described. The asymmetric ceramic hollow fibre membranes were prepared using a combined phase inversion and sintering technique. The prepared fibres were then used as support for lipase covalent immobilization in order to develop a two-separate-phase biocatalytic membrane reactor. A functionalization method was proposed in order to increase the density of the reactive hydroxyl groups on the surface of ceramic membranes, which were then amino-activated and treated with a crosslinker. The performance and the stability of the immobilized lipase were investigated as a function of the amount of the immobilized biocatalytst. Results showed that it is possible to immobilize lipase on a ceramic membrane without altering its catalytic performance (initial residual specific activity 93%), which remains constant after 6 reaction cycles.

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“…Steady‐state kinetic parameters for selected substrates are shown in Table . These studies demonstrate that DmGSTE14 indeed is catalytically competent, even though activities with the naturally occurring ITC substrates are significantly lower than the activities of the homologous GSTE6 and GSTE7 . In enzymes catalyzing several alternative reactions, it is often observed that acquisition of high activity with a particular substrate is accompanied by decreased efficiency with alternative substrates .…”

Section: Resultsmentioning

confidence: 87%

Structure and steroid isomerase activity of Drosophila glutathione transferase E14 essential for ecdysteroid biosynthesis

et al. 2020

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Ecdysteroids are critically important for the formation of the insect exoskeleton. Cholesterol is a precursor of ecdysone and its active form 20-hydroxyecdysone, but some steps in the ecdysteroid biosynthesis pathway remain unknown. An essential requirement of glutathione (GSH) transferase GSTE14 in ecdysteroid biosynthesis has been established in Drosophila melanogaster, but its function is entirely unknown. Here, we have determined the crystal structure of GSTE14 in complex with GSH and investigated the kinetic properties of GSTE14 with alternative substrates. GSTE14 has highranking steroid double-bond isomerase activity, albeit 50-fold lower than the most efficient mammalian GSTs. Corresponding steroid isomerizations are unknown in insects, and their exact physiological role remains to be shown. Nonetheless, the essential enzyme GSTE14 is here demonstrated to be catalytically competent and have a steroid-binding site.

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Glutathione transferases immobilized on nanoporous alumina: Flow system kinetics, screening, and stability

Cited by 13 publications

References 21 publications

Comparison of epsilon- and delta-class glutathioneS-transferases: the crystal structures of the glutathioneS-transferases DmGSTE6 and DmGSTE7 fromDrosophila melanogaster

Comparison of epsilon- and delta-class glutathioneS-transferases: the crystal structures of the glutathioneS-transferases DmGSTE6 and DmGSTE7 fromDrosophila melanogaster

Use of a Ceramic Membrane to Improve the Performance of Two-Separate-Phase Biocatalytic Membrane Reactor

Structure and steroid isomerase activity of Drosophila glutathione transferase E14 essential for ecdysteroid biosynthesis

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