Displaying publications 41 - 60 of 66 in total

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  1. One-pot lipase-catalyzed esterification of ε-caprolactone with methyl-d-glucopyranoside and its elongation with free 6-hydroxyhexanoate monomer units
    Saat MN, Mohamad Annuar MS
    Biotechnol Appl Biochem, 2020 May;67(3):354-365.
    PMID: 31746015 DOI: 10.1002/bab.1859
    One-pot synthesis of sugar-functionalized oligomeric caprolactone was carried out by lipase-catalyzed esterification of ε-caprolactone (ECL) with methyl-d-glucopyranoside (MGP) followed by the elongation of functionalized oligomer chain. Functionalization was performed in a custom-fabricated glass reactor equipped with Rushton turbine impeller and controlled temperature at 60 °C using tert-butanol as reaction medium. The overall reaction steps include MGP esterification of ECL monomer and its subsequent elongation by free 6-hydroxyhexanoate monomer units. A ping-pong bi-bi mechanism without ternary complex was proposed for esterification of ECL and MGP with apparent values of kinetic constant, namely maximal velocity (Vmax ), Michaelis constant for MGP (KmMGP ), and Michaelis constant for ECL (KmECL ) at 3.848 × 10-3  M H-1 , 8.189 × 10-2  M, and 6.050 M, respectively. Chain propagation step of MGP-functionalized ECL oligomer exhibits the properties of living polymerization mechanism. Linear relationship between conversion (%) and number average molecular weight, Mn (g mol-1 ), of functionalized oligomer was observed. Synthesized functionalized oligomer showed narrow range of molecular weight from 1,400 to 1,600 g mol-1 with more than 90% conversion achieved. Structural analysis confirmed the presence of covalent bond between the hydroxyl group in MGP with carboxyl end group of ECL oligomer.
    Matched MeSH terms: Biocatalysis
  2. Robust Magnetized Oil Palm Leaves Ash Nanosilica Composite as Lipase Support: Immobilization Protocol and Efficacy Study
    Onoja E, Wahab RA
    Appl Biochem Biotechnol, 2020 Oct;192(2):585-599.
    PMID: 32495234 DOI: 10.1007/s12010-020-03348-0
    Strategies to immobilize the individual enzymes are crucial for enhancing catalytic applicability and require a controlled immobilization process. Herein, protocol for immobilizing Candida rugosa lipase (CRL) onto modified magnetic silica derived from oil palm leaves ash (OPLA) was optimized for the effects of concentration of CRL, immobilization time, and temperature, monitored by titrimetric and spectrometric methods. XRD and TGA-DTG spectrometric observations indicated that OPLA-silica was well coated over magnetite (SiO2-MNPs) and CRLs were uniformly bound by covalent bonds to SiO2-MNPs (CRL/Gl-A-SiO2-MNPs). The optimized immobilization protocol showed that in the preparation of CRL/Gl-A-SiO2-MNPs, CRL with 68.3 mg/g protein loading and 74.6 U/g specific activity was achieved using 5 mg/mL of CRL, with an immobilization time of 12 h at 25 °C. The present work also demonstrated that acid-pretreated OPLA is a potential source of renewable silica, envisioning its applicability for practical use in enzymatic catalysis on solid support.
    Matched MeSH terms: Biocatalysis
  3. Recovery of value-added glycopeptides from edible bird's nest (EBN) co-products: enzymatic hydrolysis, physicochemical characteristics and bioactivity
    Ling JWA, Chang LS, Babji AS, Lim SJ
    J Sci Food Agric, 2020 Oct;100(13):4714-4722.
    PMID: 32468613 DOI: 10.1002/jsfa.10530
    BACKGROUND: Processing of edible bird's nest (EBN) requires extensive washing to remove impurities and produces huge amounts of EBN co-products, which contain mainly feathers with glycoproteins attached, which are usually discarded. This study was conducted to recover the valuable EBN glycoproteins from the waste material. Enzymatic hydrolysis was applied to recover EBN glycopeptides from EBN co-products (EBNcoP ) and processed cleaned EBN (EBNclean ) was used as control, which were then freeze-dried into EBN hydrolysates (EBNhcoP and EBNhclean , respectively).

    RESULTS: The recovery yield for EBNhclean and EBNhcoP were 89.09 ± 0.01% and 47.64 ± 0.26%, respectively, indicating nearly 50% of glycopeptide can be recovered from the waste material. Meanwhile, N-acetylneuraminic acid, a major acid sugar in EBN glycoproteins, of EBNhcoP increased by 229% from 58.6 ± 3.9 to 192.9 ± 3.1 g kg-1 , indicating the enzymatic hydrolysis removed impurities and thus enhanced the N-acetylneuraminic acid content. Total soluble protein was more than 330 g kg-1 for all the samples. Colour parameter showed that hydrolysate samples have greater L* (lightness) values. Chroma result indicates the intensity of all the samples were low (

    Matched MeSH terms: Biocatalysis
  4. Improvement of enzymatic bioxylitol production from sawdust hemicellulose: optimization of parameters
    Rafiqul ISM, Mimi Sakinah AM, Zularisam AW
    Prep Biochem Biotechnol, 2021;51(10):1060-1070.
    PMID: 33724897 DOI: 10.1080/10826068.2021.1897840
    Enzymatic production of bioxylitol from lignocellulosic biomass (LCB) provides a promising alternative to both chemical and fermentative routes. This study aimed to assess the impacts of catalytic variables on bioxylitol production from wood sawdust using xylose reductase (XR) enzyme and to optimize the bioprocess. Enzyme-based xylitol production was carried out in batch cultivation under various experimental conditions to obtain maximum xylitol yield and productivity. The response surface methodology (RSM) was followed to fine-tune the most significant variables such as reaction time, temperature, and pH, which influence the synthesis of bioxylitol from sawdust hydrolysate and to optimize them. The optimum time, temperature, and pH became were 12.25 h, 35 °C, and 6.5, respectively, with initial xylose 18.8 g/L, NADPH 2.83 g/L, XR 0.027 U/mg, and agitation 100 rpm. The maximum xylitol production was attained at 16.28 g/L with a yield and productivity of 86.6% (w/w) and 1.33 g/L·h, respectively. Optimization of catalytic parameters using sequential strategies resulted in 1.55-fold improvement in overall xylitol production. This study explores a novel strategy for using sawdust hemicellulose in bioxylitol production by enzyme technology.
    Matched MeSH terms: Biocatalysis
  5. Biocatalytic modifications of ethynodiol diacetate by fungi, anti-proliferative activity, and acetylcholineterase inhibitory of its transformed products
    Nurfazilah Wan Yusop S, Imran S, Ilham Adenan M, Ashraf K, Sultan S
    Steroids, 2021 Jul;171:108832.
    PMID: 33831403 DOI: 10.1016/j.steroids.2021.108832
    The fungal transformations of ethynodiol diacetate (1) were investigated for the first-time using Botrytis cinerea, Trichothecium roseum, and R3-2 SP 17. The metabolites obtained are as following: 17α-Ethynyl-17β-acetoxyestr-4-en-3-one-15β-ol (2), 19-nor-17a-ethynyltestosterone (3), and 17α-ethynyl-3β-hydroxy-17β-acetoxyestr-4-ene (4). The new metabolite, 2 (IC50 = 104.8 µM), which has ketone group at C-3, and the β-hydroxyl group at C-15, resulted in an almost equipotent strength with the parent compound (IC50 = 103.3 µM) against proliferation of SH-SY5Y cells. The previously reported biotransformed product, 3, showed almost equal strength to 1 against acetylcholinesterase. Molecular modelling studies were carried out to understand the observed experimental activities, and also to obtain more information on the binding mode and the interactions between the biotransformed products, and enzyme.
    Matched MeSH terms: Biocatalysis
  6. Physicochemical and functional properties of Alcalase-extracted protein hydrolysate from oil palm leaves
    Hau EH, Teh SS, Yeo SK, Mah SH
    J Sci Food Agric, 2022 Jan 15;102(1):233-240.
    PMID: 34081335 DOI: 10.1002/jsfa.11350
    BACKGROUND: The oil palm tree produces 90% of wastes and the limited usage of these wastes causes a major disposal problem in the mills. Nevertheless, these by-products have a large amount of nutritional components. Thus, the present study aimed to determine the physicochemical and functional properties of protein hydrolysates (PH) from oil palm leaves (OPL) extracted using different concentrations of Alcalase (0-10%) at 2 h of hydrolysis time.

    RESULTS: Fourier transform infrared spectral analyses showed that the enzymatic hydrolysis altered functional groups of OPL where a secondary amine was present in the PH. Changes were also observed in the thermal stability where the enthalpy heat obtained for PH (933.93-1142.57 J g-1 ) was much lower than OPL (7854.11 J g-1 ). The results showed that the PH extracted by 8% Alcalase exhibited absolute zeta potential, as well as a high emulsifying activity index (70.64 m2  g-1 of protein) and emulsion stability index (60.58 min). Furthermore, this PH showed higher solubility (96.32%) and emulsifying properties compared to other PHs. It is also comparable with commercial plant proteins, indicating that 8% Alcalase is an optimum concentration for hydrolysis.

    CONCLUSION: In summary, the physicochemical and functional properties of PH extracted from OPL showed good functional properties, suggesting that it can be used as an alternative plant protein in food industries. © 2021 Society of Chemical Industry.

    Matched MeSH terms: Biocatalysis
  7. Kinetics of enzyme-catalyzed cross-linking of feruloylated arabinan from sugar beet
    Zaidel DN, Arnous A, Holck J, Meyer AS
    J Agric Food Chem, 2011 Nov 9;59(21):11598-607.
    PMID: 21954887 DOI: 10.1021/jf203138u
    Ferulic acid (FA) groups esterified to the arabinan side chains of pectic polysaccharides can be oxidatively cross-linked in vitro by horseradish peroxidase (HRP) catalysis in the presence of hydrogen peroxide (H(2)O(2)) to form ferulic acid dehydrodimers (diFAs). The present work investigated whether the kinetics of HRP catalyzed cross-linking of FA esterified to α-(1,5)-linked arabinans are affected by the length of the arabinan chains carrying the feruloyl substitutions. The kinetics of the HRP-catalyzed cross-linking of four sets of arabinan samples from sugar beet pulp, having different molecular weights and hence different degrees of polymerization, were monitored by the disappearance of FA absorbance at 316 nm. MALDI-TOF/TOF-MS analysis confirmed that the sugar beet arabinans were feruloyl-substituted, and HPLC analysis verified that the amounts of diFAs increased when FA levels decreased as a result of the enzymatic oxidation treatment with HRP and H(2)O(2). At equimolar levels of FA (0.0025-0.05 mM) in the arabinan samples, the initial rates of the HRP-catalyzed cross-linking of the longer chain arabinans were slower than those of the shorter chain arabinans. The lower initial rates may be the result of the slower movement of larger molecules coupled with steric phenomena, making the required initial reaction of two FAs on longer chain arabinans slower than on shorter arabinans.
    Matched MeSH terms: Biocatalysis
  8. Fulltext Optimization of lipase-mediated synthesis of 1-nonene oxide using phenylacetic acid and hydrogen peroxide
    Abdulmalek E, Arumugam M, Basri M, Rahman MB
    Int J Mol Sci, 2012;13(10):13140-9.
    PMID: 23202943 DOI: 10.3390/ijms131013140
    Herein, an efficient epoxidation of 1-nonene is described. In a simple epoxidation system, commercially available Novozym 435, an immobilized Candida antarctica lipase B, and hydrogen peroxide (H(2)O(2)) were utilized to facilitate the in situ oxidation of phenylacetic acid to the corresponding peroxy acid which then reacted with 1-nonene to give 1-nonene oxide with high yield and selectivity. The aliphatic terminal alkene was epoxidised efficiently in chloroform to give an excellent yield (97%-99%) under the optimum reaction conditions, including temperature (35 °C), initial H(2)O(2) concentration (30%), H(2)O(2) amount (4.4 mmol), H(2)O(2) addition rate (one step), acid amount (8.8 mmol), and stirring speed (250 rpm). Interestingly, the enzyme was stable under the single-step addition of H(2)O(2) with a catalytic activity of 190.0 Ug-1. The entire epoxidation process was carried out within 12 h using a conventional water bath shaker.
    Matched MeSH terms: Biocatalysis
  9. Sustainable biocatalytic synthesis of L-homophenylalanine as pharmaceutical drug precursor
    Ahmad AL, Oh PC, Abd Shukor SR
    Biotechnol Adv, 2009 May-Jun;27(3):286-96.
    PMID: 19500550 DOI: 10.1016/j.biotechadv.2009.01.003
    Over the past decade, L-homophenylalanine is extensively used in the pharmaceutical industry as a precursor for production of angiotensin-converting enzyme (ACE) inhibitor, which possesses significant clinical application in the management of hypertension and congestive heart failure (CHF). A number of chemical methods have been reported thus far for the synthesis of L-homophenylalanine. However, chemical methods generally suffer from process complexity, high cost, and environmental pollution. On the other hand, enantiomerically pure L-homophenylalanine can be obtained elegantly and efficiently by employing biocatalytic methods, where it appears to be the most attractive process in terms of potential industrial applications, green chemistry and sustainability. Herein we review the biocatalytic synthesis of vital L-homophenylalanine as potentially useful intermediate in the production of pharmaceutical drugs in environmentally friendly conditions, using membrane bioreactor for sustainable biotransformation process. One envisages the future prospects of developing an integrated membrane bioreactor system with improved performance for L-homophenylalanine production.
    Matched MeSH terms: Biocatalysis
  10. Optimization of enzymatic synthesis of palm-based kojic acid ester using response surface methodology
    Ashari SE, Mohamad R, Ariff A, Basri M, Salleh AB
    J Oleo Sci, 2009;58(10):503-10.
    PMID: 19745577
    Kojic acid monooleate is a fatty acid derivative of kojic acid which can be widely used as a skin whitening agent in a cosmetic applications. In avoiding any possible harmful effects from chemically synthesized product, the enzymatic synthesis appears to be the best way to satisfy the consumer demand nowadays. The ability of immobilized lipase from Rhizomucor meihei (lipozyme RMIM) to catalyze the direct esterification of kojic acid and oleic acid was investigated. Response Surface Methodology (RSM) and 5-level-4-factor central composite rotatable were employed to evaluate the effects of synthesis parameters such as enzyme amount (0.1-0.4 g), temperature (30-60 degrees C), substrate molar ratio (1-4 mmol, kojic acid:oleic acid) and reaction time (24-48 h) on percentage molar conversion to kojic acid monooleate. Analysis of the product using TLC, GC and FTIR showed the presence of kojic acid monooleate. The optimal conditions for the enzymatic reaction were obtained after analysis with backward elimination using 0.17 g of enzyme and 4 mmol of substrate at 52.50 degrees C for 42 h. Under these conditions the esterification percentage was 37.21%. The results demonstrated that response surface methodology can be applied effectively to optimize the lipase-catalysed synthesis of kojic acid monooleate. The optimum conditions can be used to scale up the process.
    Matched MeSH terms: Biocatalysis
  11. Enzymatic synthesis of farnesyl laurate in organic solvent: initial water activity, kinetics mechanism, optimization of continuous operation using packed bed reactor and mass transfer studies
    Rahman NK, Kamaruddin AH, Uzir MH
    Bioprocess Biosyst Eng, 2011 Aug;34(6):687-99.
    PMID: 21327986 DOI: 10.1007/s00449-011-0518-y
    The influence of water activity and water content was investigated with farnesyl laurate synthesis catalyzed by Lipozyme RM IM. Lipozyme RM IM activity depended strongly on initial water activity value. The best results were achieved for a reaction medium with an initial water activity of 0.11 since it gives the best conversion value of 96.80%. The rate constants obtained in the kinetics study using Ping-Pong-Bi-Bi and Ordered-Bi-Bi mechanisms with dead-end complex inhibition of lauric acid were compared. The corresponding parameters were found to obey the Ordered-Bi-Bi mechanism with dead-end complex inhibition of lauric acid. Kinetic parameters were calculated based on this model as follows: V (max) = 5.80 mmol l(-1) min(-1) g enzyme(-1), K (m,A) = 0.70 mmol l(-1) g enzyme(-1), K (m,B) = 115.48 mmol l(-1) g enzyme(-1), K (i) = 11.25 mmol l(-1) g enzyme(-1). The optimum conditions for the esterification of farnesol with lauric acid in a continuous packed bed reactor were found as the following: 18.18 cm packed bed height and 0.9 ml/min substrate flow rate. The optimum molar conversion of lauric acid to farnesyl laurate was 98.07 ± 0.82%. The effect of mass transfer in the packed bed reactor has also been studied using two models for cases of reaction limited and mass transfer limited. A very good agreement between the mass transfer limited model and the experimental data obtained indicating that the esterification in a packed bed reactor was mass transfer limited.
    Matched MeSH terms: Biocatalysis
  12. Anticancer activity of 3-O-acylated betulinic acid derivatives obtained by enzymatic synthesis
    Ahmad FB, Ghaffari Moghaddam M, Basri M, Abdul Rahman MB
    Biosci Biotechnol Biochem, 2010;74(5):1025-9.
    PMID: 20460723
    An easy and efficient strategy to prepare betulinic acid esters with various anhydrides was used by the enzymatic synthesis method. It involves lipase-catalyzed acylation of betulinic acid with anhydrides as acylating agents in organic solvent. Lipase from Candida antarctica immobilized on an acrylic resin (Novozym 435) was employed as a biocatalyst. Several 3-O-acyl-betulinic acid derivatives were successfully obtained by this procedure. The anticancer activity of betulinic acid and its 3-O-acylated derivatives were then evaluated in vitro against human lung carcinoma (A549) and human ovarian (CAOV3) cancer cell lines. 3-O-glutaryl-betulinic acid, 3-O-acetyl-betulinic acid, and 3-O-succinyl-betulinic acid showed IC(50)<10 microg/ml against A549 cancer cell line tested and showed better cytotoxicity than betulinic acid. In an ovarian cancer cell line, all betulinic acid derivatives prepared showed weaker cytotoxicity than betulinic acid.
    Matched MeSH terms: Biocatalysis
  13. Optimized preparation and characterization of CLEA-lipase from cocoa pod husk
    Khanahmadi S, Yusof F, Amid A, Mahmod SS, Mahat MK
    J Biotechnol, 2015 May 20;202:153-61.
    PMID: 25481099 DOI: 10.1016/j.jbiotec.2014.11.015
    Cross-linked enzyme aggregate (CLEA) is easily prepared from crude enzyme and has many advantages to the environment and it is considered as an economic method in the context of industrial biocatalysis compared to free enzyme. In this work, a highly active and stable CLEA-lipase from cocoa pod husk (CPH) which is a by-product after removal of cocoa beans, were assayed for their hydrolytic activity and characterized under the optimum condition successfully. Face centered central composite design (FCCCD) under response surface methodology (RSM) was used to get the optimal conditions of the three significant factors (concentration of ammonium sulfate, concentration of glutaraldehyde and concentration of additive) to achieve higher enzyme activity of CLEA. From 20 runs, the highest activity recorded was around 9.407U (83% recovered activity) under the condition of using 20% saturated ammonium sulfate, 60mM glutaraldehyde as cross-linker and 0.17mM bovine serum albumin as feeder. Moreover, the optimal reaction temperature and pH value in enzymatic reaction for both crude enzyme and immobilized were found to be 45°C at pH 8 and 60°C at pH 8.2, respectively. A systematic study of the stability of CLEA and crude enzyme was taken with regards to temperature (25-60°C) and pH (5-10) value and in both factors, CLEA-lipase showed more stability than free lipase. The Km value of CLEA was higher compared to free enzyme (0.55mM vs. 0.08mM). The CLEA retained more than 60% of the initial activity after six cycles of reuse compared to free enzyme. The high stability and recyclability of CLEA-lipase from CPH make it efficient for different industrial applications.
    Matched MeSH terms: Biocatalysis
  14. A facile enzymatic synthesis of geranyl propionate by physically adsorbed Candida rugosa lipase onto multi-walled carbon nanotubes
    Mohamad NR, Buang NA, Mahat NA, Lok YY, Huyop F, Aboul-Enein HY, et al.
    Enzyme Microb Technol, 2015 May;72:49-55.
    PMID: 25837507 DOI: 10.1016/j.enzmictec.2015.02.007
    In view of several disadvantages as well as adverse effects associated with the use of chemical processes for producing esters, alternative techniques such as the utilization of enzymes on multi-walled carbon nanotubes (MWCNTs), have been suggested. In this study, the oxidative MWCNTs prepared using a mixture of HNO3 and H2SO4 (1:3 v/v) were used as a supportive material for the immobilization of Candida rugosa lipase (CRL) through physical adsorption process. The resulting CRL-MWCNTs biocatalysts were utilized for synthesizing geranyl propionate, an important ester for flavoring agent as well as in fragrances. Enzymatic esterification of geraniol with propionic acid was carried out using heptane as a solvent and the efficiency of CRL-MWCNTs as a biocatalyst was compared with the free CRL, considering the incubation time, temperature, molar ratio of acid:alcohol, presence of desiccant as well as its reusability. It was found that the CRL-MWCNTs resulted in a 2-fold improvement in the percentage of conversion of geranyl propionate when compared with the free CRL, demonstrating the highest yield of geranyl propionate at 6h at 55°C, molar ratio acid: alcohol of 1:5 and with the presence of 1.0g desiccant. It was evident that the CRL-MWCNTs biocatalyst could be reused for up to 6 times before a 50% reduction in catalytic efficiency was observed. Hence, it appears that the facile physical adsorption of CRL onto F-MWCNTs has improved the activity and stability of CRL as well as served as an alternative method for the synthesis of geranyl propionate.
    Matched MeSH terms: Biocatalysis
  15. Identification of functional residues essential for dehalogenation by the non-stereospecific α-haloalkanoic acid dehalogenase from Rhizobium sp. RC1
    Hamid AA, Hamid TH, Wahab RA, Huyop F
    J Basic Microbiol, 2015 Mar;55(3):324-30.
    PMID: 25727054 DOI: 10.1002/jobm.201570031
    The non-stereospecific α-haloalkanoic acid dehalogenase DehE from Rhizobium sp. RC1 catalyzes the removal of the halide from α-haloalkanoic acid D,L-stereoisomers and, by doing so, converts them into hydroxyalkanoic acid L,D-stereoisomers, respectively. DehE has been extensively studied to determine its potential to act as a bioremediation agent, but its structure/function relationship has not been characterized. For this study, we explored the functional relevance of several putative active-site amino acids by site-specific mutagenesis. Ten active-site residues were mutated individually, and the dehalogenase activity of each of the 10 resulting mutants in soluble cell lysates against D- and L-2-chloropropionic acid was assessed. Interestingly, the mutants W34→A,F37→A, and S188→A had diminished activity, suggesting that these residues are functionally relevant. Notably, the D189→N mutant had no activity, which strongly implies that it is a catalytically important residue. Given our data, we propose a dehalogenation mechanism for DehE, which is the same as that suggested for other non-stereospecific α-haloalkanoic acid dehalogenases. To the best of our knowledge, this is the first report detailing a functional aspect for DehE, and our results could help pave the way for the bioengineering of haloalkanoic acid dehalogenases with improved catalytic properties.
    Matched MeSH terms: Biocatalysis
  16. Fulltext Chemoenzymatic epoxidation of alkenes and reusability study of the phenylacetic acid
    Abdulmalek E, Arumugam M, Mizan HN, Abdul Rahman MB, Basri M, Salleh AB
    ScientificWorldJournal, 2014;2014:756418.
    PMID: 24587751 DOI: 10.1155/2014/756418
    Here, we focused on a simple enzymatic epoxidation of alkenes using lipase and phenylacetic acid. The immobilised Candida antarctica lipase B, Novozym 435 was used to catalyse the formation of peroxy acid instantly from hydrogen peroxide (H2O2) and phenylacetic acid. The peroxy phenylacetic acid generated was then utilised directly for in situ oxidation of alkenes. A variety of alkenes were oxidised with this system, resulting in 75-99% yield of the respective epoxides. On the other hand, the phenylacetic acid was recovered from the reaction media and reused for more epoxidation. Interestingly, the waste phenylacetic acid had the ability to be reused for epoxidation of the 1-nonene to 1-nonene oxide, giving an excellent yield of 90%.
    Matched MeSH terms: Biocatalysis
  17. Dendrimer-like nanoparticles based β-galactosidase assembly for enhancing its selectivity toward transgalactosylation
    Misson M, Du X, Jin B, Zhang H
    Enzyme Microb Technol, 2016 Mar;84:68-77.
    PMID: 26827776 DOI: 10.1016/j.enzmictec.2015.12.008
    Functional nanomaterials have been pursued to assemble nanobiocatalysts since they can provide unique hierarchical nanostructures and localized nanoenvironments for enhancing enzyme specificity, stability and selectivity. Functionalized dendrimer-like hierarchically porous silica nanoparticles (HPSNs) was fabricated for assembling β-galactosidase nanobiocatalysts for bioconversion of lactose to galacto-oligosaccharides (GOS). The nanocarrier was functionalized with amino (NH2) and carboxyl (COOH) groups to facilitate enzyme binding, benchmarking with non-functionalized HPSNs. Successful conjugation of the functional groups was confirmed by FTIR, TGA and zeta potential analysis. HPSNs-NH2 showed 1.8-fold and 1.1-fold higher β-galactosidase adsorption than HPSNs-COOH and HPSNs carriers, respectively, with the highest enzyme adsorption capacity of 328mg/g nanocarrier at an initial enzyme concentration of 8mg/ml. The HPSNs-NH2 and β-galactosidase assembly (HPSNs-NH2-Gal) demonstrated to maintain the highest activity at all tested enzyme concentrations and exhibited activity up to 10 continuous cycles. Importantly, HPSNs-NH2-Gal was simply recycled through centrifugation, overcoming the challenging problems of separating the nanocarrier from the reaction medium. HPSNs-NH2-Gal had distinguished catalytic reaction profiles by favoring transgalactosylation, enhancing GOS production of up to 122g/l in comparison with 56g/l by free β-galactosidase. Furthermore, it generated up to 46g/l GOS at a lower initial lactose concentration while the free counterpart had negligible GOS production as hydrolysis was overwhelmingly dominant in the reaction system. Our research findings show the amino-functionalized HPSNs can selectively promote the enzyme activity of β-galactosidase for transgalactosylation, which is beneficial for GOS production.
    Matched MeSH terms: Biocatalysis
  18. Fulltext Role of α-helical structure in organic solvent-activated homodimer of elastase strain K
    Rahman RN, Salleh AB, Basri M, Wong CF
    Int J Mol Sci, 2011;12(9):5797-814.
    PMID: 22016627 DOI: 10.3390/ijms12095797
    Recombinant elastase strain K overexpressed from E. coli KRX/pCon2(3) was purified to homogeneity by a combination of hydrophobic interaction chromatography and ion exchange chromatography, with a final yield of 48% and a 25-fold increase in specific activity. The purified protein had exhibited a first ever reported homodimer size of 65 kDa by SDS-PAGE and MALDI-TOF, a size which is totally distinct from that of typically reported 33 kDa monomer from P. aeruginosa. The organic solvent stability experiment had demonstrated a stability pattern which completely opposed the rules laid out in previous reports in which activity stability and enhancement were observed in hydrophilic organic solvents such as DMSO, methanol, ethanol and 1-propanol. The high stability and enhancement of the enzyme in hydrophilic solvents were explained from the view of alteration in secondary structures. Elastinolytic activation and stability were observed in 25 and 50% of methanol, respectively, despite slight reduction in α-helical structure caused upon the addition of the solvent. Further characterization experiments had postulated great stability and enhancement of elastase strain K in broad range of temperatures, pHs, metal ions, surfactants, denaturing agents and substrate specificity, indicating its potential application in detergent formulation.
    Matched MeSH terms: Biocatalysis/drug effects
  19. Toluene promotes lid 2 interfacial activation of cold active solvent tolerant lipase from Pseudomonas fluorescens strain AMS8
    Yaacob N, Mohamad Ali MS, Salleh AB, Rahman RNZRA, Leow ATC
    J Mol Graph Model, 2016 07;68:224-235.
    PMID: 27474867 DOI: 10.1016/j.jmgm.2016.07.003
    The utilization of cold active lipases in organic solvents proves an excellent approach for chiral synthesis and modification of fats and oil due to the inherent flexibility of lipases under low water conditions. In order to verify whether this lipase can function as a valuable synthetic catalyst, the mechanism concerning activation of the lid and interacting solvent residues in the presence of organic solvent must be well understood. A new alkaline cold-adapted lipase, AMS8, from Pseudomonas fluorescens was studied for its structural adaptation and flexibility prior to its exposure to non-polar, polar aprotic and protic solvents. Solvents such as ethanol, toluene, DMSO and 2-propanol showed to have good interactions with active sites. Asparagine (Asn) and tyrosine (Tyr) were key residues attracted to solvents because they could form hydrogen bonds. Unlike in other solvents, Phe-18, Tyr-236 and Tyr-318 were predicted to have aromatic-aromatic side-chain interactions with toluene. Non-polar solvent also was found to possess highest energy binding compared to polar solvents. Due to this circumstance, the interaction of toluene and AMS8 lipase was primarily based on hydrophobicity and molecular recognition. The molecular dynamic simulation showed that lid 2 (residues 148-167) was very flexible in toluene and Ca(2+). As a result, lid 2 moves away from the catalytic areas, leaving an opening for better substrate accessibility which promotes protein activation. Only a single lid (lid 2) showed the movement following interactions with toluene, although AMS8 lipase displayed double lids. The secondary conformation of AMS8 lipase that was affected by toluene observed a reduction of helical strands and increased coil structure. Overall, this work shows that cold active lipase, AMS8 exhibits distinguish interfacial activation and stability in the presence of polar and non-polar solvents.
    Matched MeSH terms: Biocatalysis
  20. Functional characterization of cytochrome P450 2A6 allelic variants CYP2A6*15, CYP2A6*16, CYP2A6*21, and CYP2A6*22
    Tiong KH, Yiap BC, Tan EL, Ismail R, Ong CE
    Drug Metab Dispos, 2010 May;38(5):745-51.
    PMID: 20139165 DOI: 10.1124/dmd.109.031054
    Variation in CYP2A6 levels and activity can be attributed to genetic polymorphism and, thus, functional characterization of allelic variants is necessary to define the importance of CYP2A6 polymorphism in humans. The aim of the present study was to investigate the reported alleles CYP2A6*15, CYP2A6*16, CYP2A6*21, and CYP2A6*22, in terms of the functional consequences of their mutations on the enzyme catalytic activity. With use of the wild-type CYP2A6 cDNA as template, site-directed mutagenesis was performed to introduce nucleotide changes encoding K194E substitution in CYP2A6*15, R203S substitution in CYP2A6*16, K476R substitution in CYP2A6*21, and concurrent D158E and L160I substitutions in CYP2A6*22. Upon sequence verification, the CYP2A6 wild-type and mutant constructs were individually coexpressed with NADPH-cytochrome P450 reductase in Escherichia coli. A kinetic study using a coumarin 7-hydroxylase assay indicated that CYP2A6*15 exhibited higher V(max) than the wild type, whereas all mutant constructs, except for variant CYP2A6*16, exhibited higher K(m) values. Analysis of the V(max)/K(m) ratio revealed that all mutants demonstrated 0.85- to 1.05-fold differences from the wild type, with the exception of variant CYP2A6*22, which only portrayed 39% of the wild-type intrinsic clearance. These data suggested that individuals carrying the CYP2A6*22 allele are likely to have lower metabolism of CYP2A6 substrate than individuals expressing CYP2A6*15, CYP2A6*16, CYP2A6*21, and the wild type.
    Matched MeSH terms: Biocatalysis
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