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Enzyme and Microbial Technology Aug 2023Multiple sequence alignments of three lipase isoforms from the filamentous fungus, Cordyceps militaris, have revealed that the deduced protein from their common sequence...
Multiple sequence alignments of three lipase isoforms from the filamentous fungus, Cordyceps militaris, have revealed that the deduced protein from their common sequence belongs to the Candida rugosa lipase-like group. To express the protein in its active form, recombinant lipase from C. militaris (rCML) was extra cellularly expressed in Pichia pastoris X-33 after removing its signal peptide. Purified rCML was a stable monomeric protein with a molecular mass of 90 kDa, and was highly N-mannosylated compared to the native protein (69 kDa). The catalytic efficiency (k/K) of rCML was greater than the native protein (1244.35 ± 50.88 and 1067.17 ± 29.07 mM·min, respectively), yet they had similar optimal pH values and temperatures (40 °C and pH 7.0-7.5), and showed preferences for Tween esters and short-chain triacylglycerols. Despite its monomeric conformation, interfacial activation was not observed for rCML, unlike the classical lipases. From the structural model of rCML, the binding pocket of rCML was predicted as a funnel-like structure consisting of a hollow space and an intramolecular tunnel, which is typical of C. rugosa lipase-like lipases. However, a blockage shortened the tunnel to 12-15 Å, which endows strict short-chain selectivity towards triacylglycerols and a perfect match for tricaproin (C6:0). The limited depth of the tunnel may enable accommodation of triacylglycerols with medium-to-long-chain fatty acids, which differentiates rCML from other C. rugosa lipase-like lipases with broad substrate specificities.
Topics: Lipase; Cordyceps; Recombinant Proteins; Pichia; Triglycerides; Substrate Specificity
PubMed: 37201411
DOI: 10.1016/j.enzmictec.2023.110254 -
Retinal Cases & Brief Reports Nov 2023The purpose of this study was to report a case of atypical endogenous fungal endophthalmitis caused by Candida rugosa , a rare species of nonalbicans Candida .
PURPOSE
The purpose of this study was to report a case of atypical endogenous fungal endophthalmitis caused by Candida rugosa , a rare species of nonalbicans Candida .
METHODS
This report describes a case of a 45-year-old woman who presented with a reduced visual acuity in the right eye in addition to vitreous opacity during breast cancer treatment, which was suspected as fungal endophthalmitis from medical examination and history. Various tests were performed for diagnosis.
RESULTS
Blood test results were normal, including the blood beta-D-glucan level, and blood cultures were negative. Diagnosis could not be made using systemic computed tomography and magnetic resonance imaging results. Therefore, a lesion sample was collected by using vitrectomy. C. rugosa was identified through DNA (extracted from the lesion sample) analysis using Basic Local Alignment Search Tool. The visual acuity of the right eye improved after vitrectomy.
CONCLUSION
We encountered a rare case of atypical endogenous fungal endophthalmitis caused by C. rugosa . Clinicians sometimes encounter invasive candidiasis caused by rare nonalbicans Candida species. DNA analysis using Basic Local Alignment Search Tool is effective for diagnosing such cases.
Topics: Female; Humans; Middle Aged; Candidiasis; Endophthalmitis; Vitrectomy; Eye Infections, Fungal; DNA; Antifungal Agents
PubMed: 35333842
DOI: 10.1097/ICB.0000000000001275 -
Bioresource Technology May 2024This study established a Candida rugosa lipase (CRL) system to catalyze triolein and ethyl ferulate interesterification. The products were identified, and the binding...
This study established a Candida rugosa lipase (CRL) system to catalyze triolein and ethyl ferulate interesterification. The products were identified, and the binding mode between the substrates and CRL was predicted through molecular docking. Three methods for preparing CRL-AuNPs were proposed and characterized. It was found that the addition of 40 mL of 15 nm gold nanoparticles increased the CRL activity from 3.05 U/mg to 4.75 U/mg, but the hybridization efficiency was only 32.7 %. By using 4 mL of 0.1 mg/mL chloroauric acid, the hybridization efficiency was improved to 50.7 %, but the enzyme activity was sharply decreased. However, when the molar ratio of Mb to HAuCl was 0.2, the hybridization efficiency increased to 71.8 %, and the CRL activity was also enhanced to 5.98 U/mg. Under optimal conditions, the enzyme activity of CRL-AuNPs③ was maintained at 95 % after 6 repetitions and 85.6 % after 30 days at room temperature.
Topics: Lipase; Gold; Enzymes, Immobilized; Triolein; Molecular Docking Simulation; Candida; Metal Nanoparticles; Enzyme Stability; Caffeic Acids; Saccharomycetales
PubMed: 38493938
DOI: 10.1016/j.biortech.2024.130599 -
Journal of the Science of Food and... Jun 2021In order to improve the stability of vitamin E and broaden its scope of application, an effective solution is to convert vitamin E into its derivatives. This work...
BACKGROUND
In order to improve the stability of vitamin E and broaden its scope of application, an effective solution is to convert vitamin E into its derivatives. This work developed a new Candida rugosa lipase (CRL) nanogel based on modification of ionic liquid with vinyl functional groups. This novel CRL nanogel was used in the preparation process of vitamin E succinate based on the principle of non-aqueous enzymology. At the same time, various factors including enzyme concentration, substrate molar ratio, reaction temperature and reaction time, that affect the yield of vitamin E succinate were optimized and analyzed.
RESULTS
Different solvents with various hydrophobicity parameters (LogP values) from -1.3 to 3.5 were studied, it was found that dimethyl sulfoxide (DMSO) had the lowest LogP value among organic solvents but vitamin E succinate had the highest yield in DMSO. Furthermore, the effect of different operating conditions, such as molar ratios of substrate, enzyme concentration, reaction temperature and reaction time was studied. Under the optimal process conditions (enzyme concentration 6 mg mL , substrate molar ratio 4:1, reaction temperature 55 °C and reaction time 15 h), the product yield was 62.58 ± 1.16%. CRL and CRL nanogel were characterized using Fourier-transform infrared (FTIR) spectroscopy and scanning electron microscopy (SEM). It was shown that CRL nanogel influenced the catalytic behavior of lipase significantly by changing the structure and surface properties of CRL.
CONCLUSION
Novel lipase nanogel was an excellent biocatalyst for enzymatic synthesis of vitamin E succinate. © 2020 Society of Chemical Industry.
Topics: Biocatalysis; Enzyme Stability; Enzymes, Immobilized; Esterification; Fungal Proteins; Hydrophobic and Hydrophilic Interactions; Lipase; Nanogels; Saccharomycetales; Spectroscopy, Fourier Transform Infrared; Temperature; alpha-Tocopherol
PubMed: 33216351
DOI: 10.1002/jsfa.10947 -
International Journal of Molecular... Jul 2023Green leaf volatiles (GLVs), including short chain volatile aldehydes, are widely used in the flavor and food industries because of their fresh aroma. To meet the...
Green leaf volatiles (GLVs), including short chain volatile aldehydes, are widely used in the flavor and food industries because of their fresh aroma. To meet the growing demand for natural GLVs with high added value, the use of biocatalytic processes appears as a relevant application. In such processes, vegetable oils are bioconverted into GLVs. First, the triacylglycerols of the oils are hydrolyzed by a lipase. Then, the free polyunsaturated fatty acids are converted by a lipoxygenase. Finally, volatile C6 or C9 aldehydes and 9- or 12-oxoacids are produced with a hydroperoxide lyase. Optimization of each biocatalytic step must be achieved to consider a scale-up. In this study, three oils (sunflower, hempseed, and linseed oils) and three lipases (, , and lipases) have been tested to optimize the first step of the process. The experimental design and response surface methodology (RSM) were used to determine the optimal hydrolysis conditions for each oil. Five factors were considered, i.e., pH, temperature, reaction duration, enzyme load, and oil/aqueous ratio of the reaction mixture. lipase was selected as the most efficient enzyme to achieve conversion of 96 ± 1.7%, 97.2 ± 3.8%, and 91.8 ± 3.2%, respectively, for sunflower, hempseed, and linseed oils under the defined optimized reaction conditions.
Topics: Lipase; Hydrolysis; Plant Oils; Biocatalysis; Linseed Oil; Fatty Acids, Nonesterified; Aldehydes
PubMed: 37569649
DOI: 10.3390/ijms241512274 -
International Journal of Molecular... May 2024The use of lipase immobilized on an octyl-agarose support to obtain the optically pure enantiomers of chiral drugs in reactions carried out in organic solvents is a...
The use of lipase immobilized on an octyl-agarose support to obtain the optically pure enantiomers of chiral drugs in reactions carried out in organic solvents is a great challenge for chemical and pharmaceutical sciences. Therefore, it is extremely important to develop optimal procedures to achieve a high enantioselectivity of the biocatalysts in the organic medium. Our paper describes a new approach to biocatalysis performed in an organic solvent with the use of CALB-octyl-agarose support including the application of a polypropylene reactor, an appropriate buffer for immobilization (Tris base-pH 9, 100 mM), a drying step, and then the storage of immobilized lipases in a climatic chamber or a refrigerator. An immobilized lipase B from (CALB) was used in the kinetic resolution of (,)-flurbiprofen by enantioselective esterification with methanol, reaching a high enantiomeric excess (ee = 89.6 ± 2.0%). As part of the immobilization optimization, the influence of different buffers was investigated. The effect of the reactor material and the reaction medium on the lipase activity was also studied. Moreover, the stability of the immobilized lipases: lipase from (CRL) and CALB during storage in various temperature and humidity conditions (climatic chamber and refrigerator) was tested. The application of the immobilized CALB in a polypropylene reactor allowed for receiving over 9-fold higher conversion values compared to the results achieved when conducting the reaction in a glass reactor, as well as approximately 30-fold higher conversion values in comparison with free lipase. The good stability of the CALB-octyl-agarose support was demonstrated. After 7 days of storage in a climatic chamber or refrigerator (with protection from humidity) approximately 60% higher conversion values were obtained compared to the results observed for the immobilized form that had not been stored. The new approach involving the application of the CALB-octyl-agarose support for reactions performed in organic solvents indicates a significant role of the polymer reactor material being used in achieving high catalytic activity.
Topics: Lipase; Enzymes, Immobilized; Biocatalysis; Fungal Proteins; Sepharose; Propionates; Stereoisomerism; Kinetics; Esterification; Temperature; Enzyme Stability; Candida; Solvents; Saccharomycetales
PubMed: 38791124
DOI: 10.3390/ijms25105084 -
Carbohydrate Research Jul 2020N,N,N-trimethyl chitosan (TMC), quaternized hydrophilic derivative of chitosan, has been projected to have wide applications in the pharmaceutical industry owing to its...
N,N,N-trimethyl chitosan (TMC), quaternized hydrophilic derivative of chitosan, has been projected to have wide applications in the pharmaceutical industry owing to its improved solubility at physiological conditions. However, the conventional synthesis of TMC involves toxic organic agents, which complicates its use for biological applications. Moreover, these reactions result into unwanted O-methylation and scission of the parent polymer. In the present study we have addressed these limitations by employing a green approach to synthesize TMC, by using lipase as the biocatalyst and dimethyl carbonate (DMC) as the green methylating agent, in a reaction medium comprising of ternary deep eutectic solvents (TDESs). Synthesis of TMC was carried out by using two different lipases from Burkholderia cepacia and Candida rugosa. The resulting TMC was characterized by using FTIR, H NMR, DSC, XRD. Methylation was confirmed by FTIR analysis (-CH at 1666 cm) and H NMR (?? = 3.3 ppm). DSC study revealed a lower thermal stability of TMC as compared to chitosan. These results indicated the possibility of using DMC as a green methylating agent, along with TDESs as green and sustainable solvents, for lipase catalyzed reactions. TMC was successfully synthesized and exhibited a degree of quaternization of about 12.5%, 15.69%, when synthesized used lipases from Burkholderia cepacia and Candida rugosa, respectively.
Topics: Burkholderia cepacia; Carbohydrate Conformation; Chitosan; Formates; Lipase; Saccharomycetales; Solvents
PubMed: 32505997
DOI: 10.1016/j.carres.2020.108033 -
Applied Optics Apr 2021Data acquisition and processing is a critical issue for high-speed applications, especially in three-dimensional live cell imaging and analysis. This paper focuses on...
Data acquisition and processing is a critical issue for high-speed applications, especially in three-dimensional live cell imaging and analysis. This paper focuses on sparse-data sample rotation tomographic reconstruction and analysis with several noise-reduction techniques. For the sample rotation experiments, a live Candida rugosa sample is used and controlled by holographic optical tweezers, and the transmitted complex wavefronts of the sample are recorded with digital holographic microscopy. Three different cases of sample rotation tomography were reconstructed for dense angle with a step rotation at every 2°, and for sparse angles with step rotation at every 5° and 10°. The three cases of tomographic reconstruction performance are analyzed with consideration for data processing using four noise-reduction techniques. The experimental results demonstrate potential capability in retaining the tomographic image quality, even at the sparse angle reconstructions, with the help of noise-reduction techniques.
Topics: Deep Learning; Holography; Image Processing, Computer-Assisted; Imaging, Three-Dimensional; Optical Tweezers; Rotation; Saccharomycetales; Signal-To-Noise Ratio; Tomography
PubMed: 33798139
DOI: 10.1364/AO.415284 -
Microorganisms Jan 2022Larvae of the black soldier fly (BSF) are polyphagous feeders and show tremendous bioconversion capabilities of organic matter into high-quality insect biomass....
Larvae of the black soldier fly (BSF) are polyphagous feeders and show tremendous bioconversion capabilities of organic matter into high-quality insect biomass. However, the digestion of lignocellulose-rich palm oil side streams such as palm kernel meal (PKM) is a particular challenge, as these compounds are exceptionally stable and are mainly degraded by microbes. This study aimed to investigate the suitability of BSF larvae as bioconversion agents of PKM. Since the intestinal microbiota is considered to play a key role in dietary breakdown and in increasing digestibility, the bacterial and fungal communities of BSF larvae were characterized in a culture-dependent approach and screened for their putative entomopathogenicity. The lethality of six putative candidates was investigated using intracoelomal injection. In total, 93 isolates were obtained with a bacterial share of 74% that were assigned to the four phyla , , , and . Members of the genera , and are part of the core microbiome, as they were frequently described in the gut of larvae regardless of diet, nutritional composition, or rearing conditions. With 75%, a majority of the fungal isolates belonged to the phylum . We identified several taxa already published to be able to degrade lignocelluloses, including , , yeasts, or filamentous species. The injection assays revealed pronounced differences in pathogenicity against the larvae. While caused no, weak (23.3%), moderate (53.3%), and and high (≥80%) lethality, injection resulted in 100% lethality.
PubMed: 35208774
DOI: 10.3390/microorganisms10020319 -
Applied Biochemistry and Biotechnology Nov 2022This study aimed to (i) prepare functionalized maghemite nanoparticles for immobilization of Candida rugosa lipase (CRL) by covalent binding, (ii) evaluate the...
This study aimed to (i) prepare functionalized maghemite nanoparticles for immobilization of Candida rugosa lipase (CRL) by covalent binding, (ii) evaluate the application of the immobilized derivative in the hydrolysis of waste cooking oil (WCO) to fatty acids, and (iii) assess the potential of the hydrolyzed material for biodiesel production by hydroesterification. Maghemite (γFeO) obtained by precipitation of FeCl with NHOH served as an efficient support for covalent immobilization of CRL. Fourier-transform infrared spectroscopy and hydrolytic activity analysis indicated that CRL was covalently immobilized on the surface of the maghemite support. The derivative showed an activity of 166.62 ± 8 U g in WCO hydrolysis at 40 °C and pH 6. Scanning electron microscopy revealed that, after lipase immobilization, nanoparticles became more dispersed, which is advantageous for biocatalysis reactions, as it increases the contact area with the substrate. WCO hydrolysis afforded 96 ± 0.2 wt% free fatty acids. In the second step, free fatty acids were subjected to chemical esterification with sulfuric acid, affording 94.4 ± 0.02 wt% fatty acid methyl esters (biodiesel). The findings of this study contribute to the field of biotechnology and may promote the development of enzymatic technologies for the synthesis of products of economic and social interest.
Topics: Lipase; Enzymes, Immobilized; Biofuels; Enzyme Stability; Magnetite Nanoparticles; Fatty Acids, Nonesterified; Candida; Temperature; Esterification; Biocatalysis; Sulfuric Acids
PubMed: 35789983
DOI: 10.1007/s12010-022-04046-9