Structural Adaptation of Cold-Active RTX Lipase from<i>Pseudomonas</i>sp. Strain AMS8 Revealed via Homology and Molecular Dynamics Simulation Approaches

Ali, Mohd Shukuri Mohamad; Fuzi, Siti Farhanie Mohd; Ganasen, Menega; Rahman, Raja Noor Zaliha Raja Abdul; Basri, Mahiran; Salleh, Abu Bakar

doi:10.1155/2013/925373

Cited by 23 publications

(23 citation statements)

References 32 publications

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“…Gene cloning in heterologous overexpressing hosts (Parra et al 2008;Xuezheng et al 2010;Novototskaya-Vlasova et al 2013), crystal studies (Uppenberg et al 1995;Juhl et al 2010;Durmaz et al 2013) and molecular modelling studies (Maraite et al 2013;Mohamad Ali et al 2013) strongly supports the structural attributes of cold-adapted lipases for cold habitation discussed above.…”

Section: Structural Features and Cold Adaptionmentioning

confidence: 58%

Cold active lipases – an update

Kavitha

2016

Frontiers in Life Science

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Cold active lipases (CLPs) are gaining importance nowadays as they are increasingly used in fine chemical synthesis, bioremediation, food processing and as detergent additive. These enzymes exhibit high catalytic activity at low temperatures and flexibility to act at low water medium. Since they are active at low temperatures consume less energy and also stabilize fragile compounds in the reaction medium. CLPs are commonly obtained from psychrophilic microorganisms which thrive in cold habitats. Compared to mesophilic and thermophilic lipases, only a few CLPs were studied and industrially exploited so far. CLPs (C. antarctica lipase-A and C. antarctica lipase-B) from Candida antarctica isolated from Antarctic region are the well studied and industrially employed, and many are being followed up. This review updates the CLPs reported recently and the industrial applications of CLPs. ARTICLE HISTORY

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Section: Structural Features and Cold Adaptionmentioning

confidence: 58%

Cold active lipases – an update

Kavitha

2016

Frontiers in Life Science

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“…El porcentaje de identidad entre la lipasa en estudio con la de Pseudomonas aeruginosa PAO1 fue de 45.85%, otros modelos como el desarrollado por Gupta et al (2015) utilizan 80.5% de identidad entre su lipasa y la estructura molde. Sin embargo, un porcentaje mayor no asegura el desarrollo de un buen modelo homólogo (Ali et al 2013). Así lo demuestran Messaoudi et al (2011) y Lanka et al (2015 quienes modelaron lipasas con identidades del 32% y 32.77%, respectivamente.…”

Section: Discussionunclassified

“…La estructura fue validada y sometida al análisis de acoplamiento molecular identificando dos bolsillos de interacción con los sustratos ensayados. Posteriormente, Ali et al (2013) modelaron una lipasa de Pseudomonas sp. AMS8 de 476 aa y 50 kDa, extraída de la base de datos, en este estudio, utilizaron como posibles plantillas a las lipasas de Serratia marcescens (80% de identidad) y Pseudomonas sp.…”

Section: Introductionunclassified

Determinación de secuencia y modelaje por homología de la lipasa de Marinobacter sp. LB

et al. 2018

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Las lipasas de la familia I son reconocidas a nivel industrial por sus actividades catalíticas de esterificación, interesterificación y transesterificación. En esta investigación se caracterizó por análisis in silico a la lipasa de Marinobacter sp. LB aislado de las Salinas de Pilluana, San Martín. Con tal finalidad, se amplificó el gen lip mediante la reacción en cadena de la polimerasa (PCR) de punto final y la secuencia nucleotídica fue analizada in silico. Se elucidó la estructura terciaria empleando como molde a la lipasa 1EX9 de Pseudomonas aeruginosa PAO1 y se ejecutó el acoplamiento molecular con tres sustratos. El gen lip presentó 927 pb y la proteína madura, 284 aminoácidos. La lipasa posee un peso molecular de 29.99 kDa y un pI de 8.89. Asimismo, se identificaron residuos Ser78, Asp229 e His251, típicos de la triada catalítica de una lipasa de la familia I. Además, se evidenciaron once α-hélices periféricas y siete láminas-β internas. La región del bolsillo de unión y su afinidad por lípidos fue demostrada realizando acoplamientos moleculares con trioctanoina, tributirina y trioleina, con energías de -314.28, -248.11 y -215.44 kcal/mol, respectivamente; siendo los aminoácidos de interacción Asn167, Lys106, Trp172, Thr164, Ala179. En conclusión, la estructura tridimensional de la lipasa de Marinobacter sp. LB fue construida por modelamiento homólogo y validada en base a la calidad estereoquímica y el entorno de sus aminoácidos; mientras que, los análisis de acoplamiento con sustratos de lipasas permitieron evidenciar los aminoácidos que participan en el bolsillo de unión.

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“…A varied simulation box (X = 77.49-95.05 Å, Y = 70.98-75.48 Å, Z = 66.30-66.78 Å) was filled with the toluene density appropriate to the temperature and Triton X-100 (100 molecules) was added randomly around AMS8 together with water and NaCl. The MD simulations were completed with the YASARA Structure (version 11.3.22, Krieger, Vienna, Austria) software [55,56]. Energy minimizations were performed using the steepest descent method for about~2000 steps before the simulation started with the fixed AMS8 lipase model.…”

Section: Simulation Parametersmentioning

confidence: 99%

Optimization and in Silico Analysis of a Cold-Adapted Lipase from an Antarctic Pseudomonas sp. Strain AMS8 Reaction in Triton X-100 Reverse Micelles

et al. 2018

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A moderate yield of a purified enzyme can be achieved by using the simple technique of reverse micellar extraction (RME). RME is a liquid-liquid extraction method that uses a surfactant and an organic solvent to extract biomolecules. Instead of traditional chromatographic purification methods, which are tedious and expensive, RME using the nonionic surfactant Triton X-100 and toluene is used as an alternative purification technique to purify a recombinant cold-adapted lipase, AMS8. Various process parameters were optimized to maximize the activity recovery of the AMS8 lipase. The optimal conditions were found to be 50 mM sodium phosphate buffer, pH 7, 0.125 M NaCl, and 0.07 M Triton X-100 in toluene at 10 • C. Approximately 56% of the lipase activity was successfully recovered. Structural analysis of the lipase in a reverse micelle (RM) was performed using an in silico approach. The predicted model of AMS8 lipase was simulated in the Triton X-100/toluene reverse micelles from 5 to 40 • C. The lid 2 was slightly opened at 10 • C. However, the secondary structure of AMS8 was most affected in the non-catalytic domain compared to the catalytic domain, with an increased coil conformation. These results suggest that an AMS8 lipase can be extracted using Triton X-100/water/toluene micelles at low temperature. This RME approach will be an important tool for the downstream processing of recombinant cold-adapted lipases.2 of 15 absence of co-factors [6][7][8][9]. Cold-active lipases cover a broad spectrum of biotechnological applications, such as additives in detergents (cold washing), an additive in food industries (fermentation, cheese manufacture, bakery, meat tenderizing), bioremediations (Digesters, composting, oil, or xenobiotic biology applications), and molecular biology applications [10][11][12].Several conventional purification methods for cold-adapted lipases have been applied for many years in industrial and laboratory procedures [13,14]. For example, the lipase KE38 from Pseudomonas fluorescens KE38 was successfully purified using (NH 4 ) 2 SO 4 precipitation, Sephadex G-100 and, lastly, ultrafiltration with a recovery of 54.99% and a 41.13-fold purification [15]. Likewise, three purification steps were used to purify extracellular LipP from Pseudomonas sp. strain BII-I by using 20-65% (NH 4 ) 2 SO 4 precipitation followed by a diethylaminoethyl (DEAE)-cellulose and graphite column with 38-fold purification and a recovery of 17% of the lipase activity [13]. Although a good yield of purified enzyme was obtained from these chromatographic techniques, the production and purification of large amounts of enzyme are often laborious and expensive, which hinders applications for these enzymes [16]. Therefore, a standard method that enables the purification of large amounts of highly pure, active, and stable enzymes is urgently needed. Reverse micellar extraction is one of the alternative purification methods under consideration for continuous downstream processing of bulk enzymes.A reverse micelle (RM) is a surfactant...

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Structural Adaptation of Cold-Active RTX Lipase fromPseudomonassp. Strain AMS8 Revealed via Homology and Molecular Dynamics Simulation Approaches

Cited by 23 publications

References 32 publications

Cold active lipases – an update

Cold active lipases – an update

Determinación de secuencia y modelaje por homología de la lipasa de Marinobacter sp. LB

Optimization and in Silico Analysis of a Cold-Adapted Lipase from an Antarctic Pseudomonas sp. Strain AMS8 Reaction in Triton X-100 Reverse Micelles

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