Plou FJ

References (9)

Title : Thermophilic Carboxylesterases from Hydrothermal Vents of the Volcanic Island of Ischia Active on Synthetic and Biobased Polymers and Mycotoxins - Distaso_2023_Appl.Environ.Microbiol__e0170422
Author(s) : Distaso MA , Chernikova TN , Bargiela R , Coscolin C , Stogios P , Gonzalez-Alfonso JL , Lemak S , Khusnutdinova AN , Plou FJ , Evdokimova E , Savchenko A , Lunev EA , Yakimov MM , Golyshina OV , Ferrer M , Yakunin AF , Golyshin PN
Ref : Applied Environmental Microbiology , :e0170422 , 2023
Abstract : Hydrothermal vents are geographically widespread and host microorganisms with robust enzymes useful in various industrial applications. We examined microbial communities and carboxylesterases of two terrestrial hydrothermal vents of the volcanic island of Ischia (Italy) predominantly composed of Firmicutes, Proteobacteria, and Bacteroidota. High-temperature enrichment cultures with the polyester plastics polyhydroxybutyrate and polylactic acid (PLA) resulted in an increase of Thermus and Geobacillus species and to some extent Fontimonas and Schleiferia species. The screening at 37 to 70 degreesC of metagenomic fosmid libraries from above enrichment cultures identified three hydrolases (IS10, IS11, and IS12), all derived from yet-uncultured Chloroflexota and showing low sequence identity (33 to 56%) to characterized enzymes. Enzymes expressed in Escherichia coli exhibited maximal esterase activity at 70 to 90 degreesC, with IS11 showing the highest thermostability (90% activity after 20-min incubation at 80 degreesC). IS10 and IS12 were highly substrate promiscuous and hydrolyzed all 51 monoester substrates tested. Enzymes were active with PLA, polyethylene terephthalate model substrate, and mycotoxin T-2 (IS12). IS10 and IS12 had a classical alpha/beta-hydrolase core domain with a serine hydrolase catalytic triad (Ser155, His280, and Asp250) in their hydrophobic active sites. The crystal structure of IS11 resolved at 2.92 A revealed the presence of a N-terminal beta-lactamase-like domain and C-terminal lipocalin domain. The catalytic cleft of IS11 included catalytic Ser68, Lys71, Tyr160, and Asn162, whereas the lipocalin domain enclosed the catalytic cleft like a lid and contributed to substrate binding. Our study identified novel thermotolerant carboxylesterases with a broad substrate range, including polyesters and mycotoxins, for potential applications in biotechnology. IMPORTANCE High-temperature-active microbial enzymes are important biocatalysts for many industrial applications, including recycling of synthetic and biobased polyesters increasingly used in textiles, fibers, coatings and adhesives. Here, we identified three novel thermotolerant carboxylesterases (IS10, IS11, and IS12) from high-temperature enrichment cultures from Ischia hydrothermal vents and incubated with biobased polymers. The identified metagenomic enzymes originated from uncultured Chloroflexota and showed low sequence similarity to known carboxylesterases. Active sites of IS10 and IS12 had the largest effective volumes among the characterized prokaryotic carboxylesterases and exhibited high substrate promiscuity, including hydrolysis of polyesters and mycotoxin T-2 (IS12). Though less promiscuous than IS10 and IS12, IS11 had a higher thermostability with a high temperature optimum (80 to 90 degreesC) for activity and hydrolyzed polyesters, and its crystal structure revealed an unusual lipocalin domain likely involved in substrate binding. The polyesterase activity of these enzymes makes them attractive candidates for further optimization and potential application in plastics recycling.
ESTHER : Distaso_2023_Appl.Environ.Microbiol__e0170422
PubMedSearch : Distaso_2023_Appl.Environ.Microbiol__e0170422
PubMedID: 36719236
Gene_locus related to this paper: 9bact-estC55.8n1 , 9bact-IS10

Title : Transforming an esterase into an enantioselective catecholase through bioconjugation of a versatile metal-chelating inhibitor - Fernandez-Lopez_2023_Chem.Commun.(Camb)__
Author(s) : Fernandez-Lopez L , Cea-Rama I , Alvarez-Malmagro J , Ressmann AK , Gonzalez-Alfonso JL , Coscolin C , Shahgaldian P , Plou FJ , Modregger J , Pita M , Sanz-Aparicio J , Ferrer M
Ref : Chem Commun (Camb) , : , 2023
Abstract : Metal complexes introduced into protein scaffolds can generate versatile biomimetic catalysts endowed with a variety of catalytic properties. Here, we synthesized and covalently bound a bipyridinyl derivative to the active centre of an esterase to generate a biomimetic catalyst that shows catecholase activity and enantioselective catalytic oxidation of (+)-catechin.
ESTHER : Fernandez-Lopez_2023_Chem.Commun.(Camb)__
PubMedSearch : Fernandez-Lopez_2023_Chem.Commun.(Camb)__
PubMedID: 37376994
Gene_locus related to this paper: 9zzzz-a0a2k8jn75

Title : Crystal structure of a family VIII beta-lactamase fold hydrolase reveals the molecular mechanism for its broad substrate scope - Cea-Rama_2022_FEBS.J__
Author(s) : Cea-Rama I , Coscolin C , Gonzalez-Alfonso JL , Raj J , Vasiljevic M , Plou FJ , Ferrer M , Sanz-Aparicio J
Ref : Febs J , : , 2022
Abstract : Family VIII esterases present similarities to class C beta-lactamases, which show nucleophilic serines located at the S-X-X-K motif instead of the G-X-S-X-G or G-D-S-(L) motif shown by other carboxylesterase families. Here, we report the crystal structure of a novel family VIII (subfamily VIII. I) esterase (EH(7) ; denaturing temperature, 52.6+/-0.3 degreesC; pH optimum 7.0-9.0) to deepen its broad substrate range. Indeed, the analysis of the substrate specificity revealed its capacity to hydrolyse nitrocefin as a model chromogenic cephalosporin substrate (40.4 +/- 11.4 units/g), as well as a large battery of 66 structurally different esters (up to 1730 min(-1) ), including bis(2-hydroxyethyl)-terephthalate (241.7 +/- 8.5 units/g) and the mycotoxin T-2 (1220 +/- 52 units/g). It also showed acyltransferase activity through the synthesis of benzyl 3-oxobutanoate (40.4 +/- 11.4 units/g) from benzyl alcohol and vinyl acetoacetate. Such a broad substrate scope is rare among family VIII esterases and lipolytic enzymes. Structural analyses of free and substrate-bound forms of this homo-octamer esterase suggest that EH(7) presents a more opened and exposed S1 site having no steric hindrance for the entrance of substrates to the active site, more flexible R1, R2 and R3 regions allowing the binding of a wide spectrum of substrates into the active site, as well as small residues in the conserved motif Y-X-X containing the catalytic Tyr enabling the entrance of large substrates. These unique structural elements in combination with docking experiments allowed us to gain valuable insights into the substrate specificity of this esterase and possible others belonging to family VIII.
ESTHER : Cea-Rama_2022_FEBS.J__
PubMedSearch : Cea-Rama_2022_FEBS.J__
PubMedID: 35694902

Title : Genetically engineered proteins with two active sites for enhanced biocatalysis and synergistic chemo- and biocatalysis - Alonso_2020_Nat.Catal_3_319
Author(s) : Alonso S , Santiago G , Cea-Rama I , Fernandez-Lopez L , Coscolin C , Modregger J , Ressmann A , Martinez-Martinez M , Marrero H , Bargiela R , Pita M , Gonzalez-Alfonso JL , Briand M , Rojo D , Barbas C , Plou FJ , Golyshin PN , Shahgaldian P , Sanz-Aparicio J , Guallar V , Ferrer M
Ref : Nature Catalysis , 3 :319 , 2019
Abstract : Enzyme engineering has allowed not only the de novo creation of active sites catalysing known biological reactions with rates close to diffusion limits, but also the generation of abiological sites performing new-to-nature reactions. However, the catalytic advantages of engineering multiple active sites into a single protein scaffold are yet to be established. Here, we report on pro-teins with two active sites of biological and/or abiological origin, for improved natural and non-natural catalysis. The approach increased the catalytic properties, such as enzyme efficiency, substrate scope, stereoselectivity and optimal temperature window, of an esterase containing two biological sites. Then, one of the active sites was metamorphosed into a metal-complex chemocatalytic site for oxidation and Friedel-Crafts alkylation reactions, facilitating synergistic chemo- and biocatalysis in a single protein. The transformations of 1-naphthyl acetate into 1,4-naphthoquinone (conversion approx. 100%) and vinyl crotonate and benzene into 3-phenylbutyric acid (>=83%; e.e. >99.9%) were achieved in one pot with this artificial multifunc-tional metalloenzyme.
ESTHER : Alonso_2020_Nat.Catal_3_319
PubMedSearch : Alonso_2020_Nat.Catal_3_319
Gene_locus related to this paper: 9bact-LAE6

Title : Synthesis and emulsifying properties of carbohydrate fatty acid esters produced from Agave tequilana fructans by enzymatic acylation - Casas-Godoy_2016_Food.Chem_204_437
Author(s) : Casas-Godoy L , Arrizon J , Arrieta-Baez D , Plou FJ , Sandoval G
Ref : Food Chem , 204 :437 , 2016
Abstract : Carbohydrate fatty acid esters are non-ionic surfactants with a broad spectrum of applications. These molecules are generally synthesized using short carbohydrates or linear fructans; however in this research carbohydrate fatty acid esters were produced for the first time with branched fructans from Agave tequilana. Using immobilized lipases we successfully acylated A. tequilana fructans with vinyl laurate, obtaining products with different degrees of polymerization (DP). Lipozyme 435 was the most efficient lipase to catalyze the transesterification reaction. HPLC and ESI-MS analysis proved the presence of a mixture of acylated products as a result of the chemical complexity of fructans in the A. tequilana. The ESI-MS spectra showed a molecular mass shift between 183 and 366g/mol for fructooligosaccharides with a DP lower than 6, which indicated the presence of Agave fructans that had been mono- and diacylated with lauric acid. The carbohydrate fatty acid esters (CFAE) obtained showed good emulsifying properties in W/O emulsions.
ESTHER : Casas-Godoy_2016_Food.Chem_204_437
PubMedSearch : Casas-Godoy_2016_Food.Chem_204_437
PubMedID: 26988522

Title : Lipase-catalyzed modification of phenolic antioxidants - Torres_2012_Methods.Mol.Biol_861_435
Author(s) : Torres P , Reyes-Duarte D , Ballesteros A , Plou FJ
Ref : Methods Mol Biol , 861 :435 , 2012
Abstract : The chemical acylation of natural antioxidants may improve their oxidative and thermal stability, as well as modify their hydrophile-lipophile balance (HLB). These processes are generally carried out under harsh conditions using strongly corrosive acids. In contrast, lipase-catalyzed acylation is characterized by mild reaction conditions, low energy requirements, and a minimization of side reactions. We report the one-step enzymatic acylation of a phenolic antioxidant (alpha-tocopherol) and a polyphenol (resveratrol) by lipase-catalyzed transesterification. In particular, the regioselectivity of resveratrol acylation can be controlled by an adequate selection of the biocatalyst.
ESTHER : Torres_2012_Methods.Mol.Biol_861_435
PubMedSearch : Torres_2012_Methods.Mol.Biol_861_435
PubMedID: 22426732

Title : Regioselective lipase-catalyzed synthesis of 3-o-acyl derivatives of resveratrol and study of their antioxidant properties - Torres_2010_J.Agric.Food.Chem_58_807
Author(s) : Torres P , Poveda A , Jimenez-Barbero J , Ballesteros A , Plou FJ
Ref : Journal of Agricultural and Food Chemistry , 58 :807 , 2010
Abstract : One of the approaches to increasing the bioavailability of resveratrol is to protect its 3-OH phenolic group. In this work, regioselective acylation of resveratrol at 3-OH was achieved by transesterification with vinyl acetate catalyzed by immobilized lipase from Alcaligenes sp. (lipase QLG). The maximum yield of 3-O-acetylresveratrol was approximately 75%, as the lipase also catalyzes its further acetylation affording the diester 3,4'-di-O-acetylresveratrol and finally the peracetylated derivative. Long saturated and unsaturated fatty acid vinyl esters were also effective as acyl donors with similar regioselectivity. In contrast, lipase B from Candida antarctica catalyzes the acylation of the phenolic group 4'-OH with 80% yield and negligible formation of higher esters. The analysis of the antioxidant properties showed that the Trolox equivalent antioxidant capability (TEAC) values for the acetyl and stearoyl derivatives at 3-OH were, respectively, 40% and 25% referred to resveratrol. The addition of an acyl chain in the 3-OH position caused a higher loss of activity compared with that at the 4'-OH.
ESTHER : Torres_2010_J.Agric.Food.Chem_58_807
PubMedSearch : Torres_2010_J.Agric.Food.Chem_58_807
PubMedID: 20017485

Title : Conversion of a carboxylesterase into a triacylglycerol lipase by a random mutation -
Author(s) : Reyes-Duarte D , Polaina J , Lopez-Cortes N , Alcalde M , Plou FJ , Elborough K , Ballesteros A , Timmis KN , Golyshin PN , Ferrer M
Ref : Angew Chem Int Ed Engl , 44 :7553 , 2005
PubMedID: 16254934

Title : Production, isolation and characterization of a sterol esterase from Ophiostoma piceae - Calero-Rueda_2002_Biochim.Biophys.Acta_1599_28
Author(s) : Calero-Rueda O , Plou FJ , Ballesteros A , Martinez AT , Martinez MJ
Ref : Biochimica & Biophysica Acta , 1599 :28 , 2002
Abstract : We studied extracellular sterol esterase production by the ascomycete Ophiostoma piceae in liquid culture. Esterase activity was found in low levels in glucose medium but it was strongly induced by olive oil. An esterase was purified from the 0.5% olive oil-supplemented cultures using ultrafiltration followed by a single chromatographic step on a hydrophobic interaction column. The enzyme was a glycoprotein with 8% N-linked carbohydrate content, a molecular mass by SDS/PAGE around 56.5 kDa and an isoelectric point of 3.3. Its N-terminal sequence was TTVNVKYPEGEVV. Substrate specificity studies showed that the O. piceae esterase hydrolyzes p-nitrophenol esters, tributyrin, triolein and different cholesterol esters. Both affinity (Km) and catalytic constant (k(cat)) were positively affected by the length of the fatty acid esterifying glycerol and cholesterol. The presence of double bonds in the acyl chain increased the enzyme efficiency, although it affected the k(cat) values rather than the Km on the cholesterol esters. The O. piceae enzyme showed no interfacial activation. This enzyme could have biotechnological applications in paper manufacturing since it efficiently hydrolyzes both triglycerides and sterol esters, which form pitch deposits during manufacturing of softwood and hardwood paper pulps, respectively.
ESTHER : Calero-Rueda_2002_Biochim.Biophys.Acta_1599_28
PubMedSearch : Calero-Rueda_2002_Biochim.Biophys.Acta_1599_28
PubMedID: 12479402
Gene_locus related to this paper: 9pezi-q2tfw1