A novel thermoalkalostable esterase from Acidicaldus sp. strain USBA-GBX-499 with enantioselectivity isolated from an acidic hot springs of Colombian Andes

López, Gina; Chow, Jennifer; Bongen, Patrick; Lauinger, Benjamin; Pietruszka, Jörg; Streit, Wolfgang R.; Baena, Sandra

doi:10.1007/s00253-014-5775-7

Cited by 29 publications

(20 citation statements)

References 69 publications

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“…Finally, addition of the serine hydrolase inhibitor phenylmethylsulfonyl fluoride (PMSF) resulted in a dramatic decrease in EstDZ3 activity, thus indicating that a serine residue is involved in the catalytic mechanism of this new enzyme ( Table 2 ). These levels of tolerance against the presence of metals and detergents are typical for thermostable enzymes (Peng et al, 2011; López et al, 2014). …”

Section: Introductionmentioning

confidence: 99%

EstDZ3: A New Esterolytic Enzyme Exhibiting Remarkable Thermostability

et al. 2016

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Lipolytic enzymes that retain high levels of catalytic activity when exposed to a variety of denaturing conditions are of high importance for a number of biotechnological applications. In this study, we aimed to identify new lipolytic enzymes, which are highly resistant to prolonged exposure to elevated temperatures. To achieve this, we searched for genes encoding for such proteins in the genomes of a microbial consortium residing in a hot spring located in China. After performing functional genomic screening on a bacterium of the genus Dictyoglomus, which was isolated from this hot spring following in situ enrichment, we identified a new esterolytic enzyme, termed EstDZ3. Detailed biochemical characterization of the recombinant enzyme, revealed that it constitutes a slightly alkalophilic and highly active esterase against esters of fatty acids with short to medium chain lengths. Importantly, EstDZ3 exhibits remarkable thermostability, as it retains high levels of catalytic activity after exposure to temperatures as high as 95°C for several hours. Furthermore, it exhibits very good stability against exposure to high concentrations of a variety of organic solvents. Interestingly, EstDZ3 was found to have very little similarity to previously characterized esterolytic enzymes. Computational modeling of the three-dimensional structure of this new enzyme predicted that it exhibits a typical α/β hydrolase fold that seems to include a “subdomain insertion”, which is similar to the one present in its closest homolog of known function and structure, the cinnamoyl esterase Lj0536 from Lactobacillus johnsonii. As it was found in the case of Lj0536, this structural feature is expected to be an important determinant of the catalytic properties of EstDZ3. The high levels of esterolytic activity of EstDZ3, combined with its remarkable thermostability and good stability against a range of organic solvents and other denaturing agents, render this new enzyme a candidate biocatalyst for high-temperature biotechnological applications.

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Section: Introductionmentioning

confidence: 99%

EstDZ3: A New Esterolytic Enzyme Exhibiting Remarkable Thermostability

et al. 2016

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“…In contrast, phenotypic screening approaches apply activity-based assays enabling the discovery of so far unknown proteins belonging to completely novel families. Since the environmental conditions shape the microbial diversity, the choice of the respective habitat is essential for successful mining for novel biocatalysts as shown for cold, hot, and halophilic, habitats56. Furthermore, nutrient availability largely determines the spectrum of enzymes to be identified78.…”

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confidence: 99%

Metagenomic discovery of novel enzymes and biosurfactants in a slaughterhouse biofilm microbial community

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Rausch

Kovačić

et al. 2016

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DNA derived from environmental samples is a rich source of novel bioactive molecules. The choice of the habitat to be sampled predefines the properties of the biomolecules to be discovered due to the physiological adaptation of the microbial community to the prevailing environmental conditions. We have constructed a metagenomic library in Escherichia coli DH10b with environmental DNA (eDNA) isolated from the microbial community of a slaughterhouse drain biofilm consisting mainly of species from the family Flavobacteriaceae. By functional screening of this library we have identified several lipases, proteases and two clones (SA343 and SA354) with biosurfactant and hemolytic activities. Sequence analysis of the respective eDNA fragments and subsequent structure homology modelling identified genes encoding putative N-acyl amino acid synthases with a unique two-domain organisation. The produced biosurfactants were identified by NMR spectroscopy as N-acyltyrosines with N-myristoyltyrosine as the predominant species. Critical micelle concentration and reduction of surface tension were similar to those of chemically synthesised N-myristoyltyrosine. Furthermore, we showed that the newly isolated N-acyltyrosines exhibit antibiotic activity against various bacteria. This is the first report describing the successful application of functional high-throughput screening assays for the identification of biosurfactant producing clones within a metagenomic library.

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“…This finding is contrary to those previously reported on lipolytic enzymes, Universitas Scientiarum Vol. 22 (1): http://ciencias.javeriana.edu.co/investigacion/universitas-scientiarum which showed that metallic ions can play an important role in maintaining enzymatic structural stability and integrity [50], for example the presence of Fe 3+ and Mn 2+ at 1 mM increased the relative enzymatic activity of the 499EST esterase of Acidicaldus USBA-GBX-499 [23]. Moreover, under particular conditions, the presence of metallic cofactors facilitated the correct folding of an enzyme [51] and stabilized the structure of the catalytic site of Geobacillus zalihae lipase [52].…”

Section: Effect Of Metallic Ions On the Activity Of The Enzyme 505 Lipmentioning

confidence: 99%

“…The enzymatic reaction was performed under the following conditions: 425 µl of Tris-HCl buffer (50 mM, pH 7.5), 25 µl of the substrate pNPester at a final concentration of 0.5 mM and 25 µl of the sample [23]. The absorbance at 405 nm was measured using a HACH DR 5000 ™ UV-Vis Spectrophotometer (HACH Loveland, CO, U.S.A.).…”

Section: Evaluation Of the Intracellular Lipolytic Activity Using P-nmentioning

confidence: 99%

“…Other lipolytic enzymes derived from acidophilic organisms include the lipolytic enzyme produced by the thermoacidophilic microorganism Alicyclobacillus acidocaldarius, with an optimal temperature of 60 ºC [48], and the thermophilic esterase produced by Acidicaldus sp. USBA-GBX-499, with an optimal temperature of 55 ºC [23].…”

Section: Partial Purification Of the Lipolytic Enzyme Of A Facilis Umentioning

confidence: 99%

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Response Surface Methodology (RSM) for analysing culture conditions of Acidocella facilis strain USBA-GBX-505 and Partial Purification and Biochemical Characterization of Lipase 505 LIP

et al. 2017

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Using Response Surface Methodology (RSM) we evaluated the culture conditions (nitrogen source, carbon source, pH and agitation rate) that increase the biomass of Acidocella facilis strain USBA-GBX-505 and therefore enhance the production of its lipolytic enzyme, 505 LIP. RSM results revealed that yeast extract and agitation were key culture factors that increased the growth-associated lipolytic activity by 4.5-fold (from 0.13 U.mg -1 to 0.6 U.mg -1). The 505 LIP lipase was partially purified using size-exclusion chromatography and ion-exchange chromatography. Its molecular weight was >77 kDa. The enzyme shows its optimum catalytic activity at 55 °C and pH 7.5. EDTA, PMSF, 1-butanol and DMSO inhibited enzymatic activity, whereas Tween 20, acetone, glycerol and methanol increased it. Metallic ions are not required for the activity of 505 LIP, and even have an inhibitory effect on the enzyme. This study shows the potential use of A. facilis strain USBA-GBX-505 for the production of a newly identified lipolytic enzyme, 505 LIP, which is stable at moderate temperatures and in the presence of organic solvents. These are important characteristics for the synthesis of many useful products.

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A novel thermoalkalostable esterase from Acidicaldus sp. strain USBA-GBX-499 with enantioselectivity isolated from an acidic hot springs of Colombian Andes

Cited by 29 publications

References 69 publications

EstDZ3: A New Esterolytic Enzyme Exhibiting Remarkable Thermostability

EstDZ3: A New Esterolytic Enzyme Exhibiting Remarkable Thermostability

Metagenomic discovery of novel enzymes and biosurfactants in a slaughterhouse biofilm microbial community

Response Surface Methodology (RSM) for analysing culture conditions of Acidocella facilis strain USBA-GBX-505 and Partial Purification and Biochemical Characterization of Lipase 505 LIP

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