Anatomy of lipase binding sites: the scissile fatty acid binding site

Pleiss, Jürgen; Fischer, Markus; Schmid, Rolf D.

doi:10.1016/s0009-3084(98)00030-9

Cited by 433 publications

(318 citation statements)

References 40 publications

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“…25 In addition, no correspondence was found between the hydrolytic activities (Table 1) Therefore, some insight into the enzyme mechanism may be gained by considering a comparative analysis of the structures described for some lipases used in this work. According to Pleiss et al, 27 lipases have a long, hydrophobic scissile fatty acid binding site located inside the binding pocket on top of the central β sheet (in a crevice, on the wall of a binding funnel or in a tunnel). The protein surface on the border of this pocket is hydrophobic and it is supposed to interact with the hydrophobic substrate interface.…”

Section: Influence Of Lipase Sourcementioning

confidence: 99%

“…Both lipases are active in substrates containing medium chain fatty acids such as lauric acid, though the Rhizomucor miehei lipase has its active catalytic triad on its surface, in a long and well defined hydrophobic crevice, in contrast to Candida antartica B lipase, which has a small hydrophobic area located on the wall of the binding funnel. 27 For Candida rugosa lipase, that has an aliphatic site positioned in a long tunnel inside the protein with a wide entrance involved in the binding of the scissile fatty acid chain, 29 previous studies showed a clear preference for short and medium chain fatty acids, mainly C 4 , C 8 and C 10 . 30 This could explain the poor molar conversion attained by this lipase source under the reaction conditions used here.…”

Section: Influence Of Lipase Sourcementioning

confidence: 99%

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Enzymatic synthesis of glyceride esters in solvent-free system: influence of the molar ratio, lipase source and functional activating agent of the support

Freitas¹,

Pérez²,

Santos³

et al. 2007

J. Braz. Chem. Soc.

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Este trabalho avaliou a influência de fatores que afetam a síntese de ésteres glicéricos, tais como: razão molar entre os reagentes, fonte de lipase e tipo de agente de ativação do suporte obtido pela técnica sol-gel. Foram empregadas preparações comerciais de lipases imobilizadas em partículas de polissiloxano-álcool polivinílico (POS-PVA) previamente ativadas com diferentes agentes (glutaraldeído, metaperiodato de sódio e carbonildiimidazol). Esses resultados foram comparados com aqueles obtidos por preparações de lipase imobilizada disponíveis comercialmente (Lipozyme IM 20 , Novozym 435, Lipozyme RM IM e Lipozyme TL IM). O meio reacional contendo excesso de glicerol favoreceu a síntese de ésteres glicéricos e a Lipozyme IM 20 foi a preparação de lipase imobilizada mais adequada para a síntese, atingindo conversões molares maiores que 94%. O uso da lipase CALB L imobilizada em POS-PVA também alcançou resultados satisfatórios (cerca de 80% de conversão) possibilitando a formação de 36% de 2,3-diidroxipropil dodecanoato (monolaurina).This work assessed the influence of important factors that affect the synthesis of glyceride esters in solvent-free systems, such as: glycerol/fatty acid molar ratio, lipase source and activating agent of the support obtained by the sol-gel technique. Commercial lipase preparations were immobilized on polysiloxane-polyvinyl alcohol particles (POS-PVA) previously activated with different agents (glutaraldehyde, sodium metaperiodate and carbonyldiimidazole) and their performance on the esterification reaction was compared with commercial preparations of immobilized lipase (Lipozyme IM 20 , Novozym 435, Lipozyme RM IM and Lipozyme TL IM). The reaction medium containing excess glycerol favored the glyceride ester synthesis and the Lipozyme IM 20 was found to be the most suitable immobilized lipase preparation, attaining molar conversions higher than 94%. The use of CAL B Lipase immobilized on POS-PVA also provided satisfactory performance (conversion of about 80%) and allowed the formation of 36% wt of 2,3-dihydroxypropyl dodecanoate (monolaurin).Keywords: glycerides, lipase, esterification, hybrid support, immobilization IntroductionThe demand of the industries to operate their processes under sustained conditions such as green chemistry or clean technology systems is becoming an important issue nowadays and thus requests the availability of tailored biocatalysts to operate under these conditions. 1 Therefore, the need for developing active and stable immobilized biocatalysts for subsequent use in biotransformations has been highlighted. 2 Among the most important biocatalysts carrying out novel reactions in both aqueous and non aqueous media, lipases stand out due to their ability to utilize a wide spectrum of substrates, high stability towards extremes of temperature, pH and organic solvents, and chemo-, regio-and enantioselectivity. [3][4][5] Although most enzymatic syntheses are performed in presence of organic non-polar solvents, 6 the elimination of these compounds is technicall...

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Section: Influence Of Lipase Sourcementioning

confidence: 99%

Section: Influence Of Lipase Sourcementioning

confidence: 99%

Enzymatic synthesis of glyceride esters in solvent-free system: influence of the molar ratio, lipase source and functional activating agent of the support

Freitas¹,

Pérez²,

Santos³

et al. 2007

J. Braz. Chem. Soc.

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“…Na reação dos ácidos palmítico (C16) e esteárico (C18) com npentanol, as conversões foram um pouco menores que a obtida para o ácido mirístico (66%), sendo de 62 e 52%, respectivamente. Devese considerar a influência dos efeitos estéreos causados pelo aumento da cadeia alquílica, dificultando a formação do complexo acil-enzima, que é a primeira etapa do mecanismo proposto para as reações catalisadas por lipases 47 .…”

Section: Influência Do Comprimento Das Cadeias Alquílicas Do áCido E unclassified

Imobilização de lipases em filme de caseinato de sódio/glicerol: aplicação na síntese de ésteres

Sebrão¹,

Silva²,

Nascimento³

et al. 2007

Quím. Nova

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Recebido em 12/7/06; aceito em 1/12/06; publicado na web em 24/7/07 LIPASE IMMOBILIZATION IN SODIUM CASEINATE/GLYCEROL FILM: APPLICATION IN ESTER SYNTHESIS. Lipases from different sources were immobilized in sodium caseinate/glycerol film and used in the esterification reactions of aliphatic acids with alcohols in the presence of organic solvents. Lipases from Pseudomonas sp and Rhizopus oryzae were selected and the influence of several parameters was analyzed, including: lipase loading, organic solvent polarity, reaction temperature, chain length of alcohol and acid and enzyme/support reuse. For comparison, free enzymes were used under similar experimental conditions. Keywords: sodium caseinate/glycerol; lipases; immobilization. INTRODUÇÃOAs enzimas apresentam várias propriedades que as tornam atrativas como catalisadores para biotransformações, destacando-se o uso na indústria químico-farmacêutica. Estas proteínas são catalisadores versáteis, existindo em geral um processo enzimático equivalente para cada tipo de reação orgânica [1][2][3] .A partir da década 80 tem sido observado um interesse crescente no desenvolvimento de técnicas de imobilização de enzimas e/ou microrganismos, visando minimizar os efeitos causados pelo seu uso em ambientes adversos, tais como solventes orgânicos, variações no pH e/ou altas temperaturas 1,4 . A imobilização do biocatalisador em um suporte, sem prejuízo de sua atividade por um período razoável de tempo, pode assegurar sua repetida utilização ou mesmo o uso em reatores contínuos, resultando em economia nos processos industriais. Assim, de modo geral, a utilização de materiais imobilizados além de diminuir o custo por análise, aumenta a rapidez e a exatidão do processo [4][5][6] .Um dos fatores que sempre deve ser considerado na utilização de um determinado sistema de imobilização é o tipo de interação entre o suporte e o biocatalisador, que pode influenciar diretamente na estabilidade e nos efeitos cinéticos da catálise 1,4-9 . Os processos de imobilização podem ocorrer por adsorção ou ligação da enzima em um material insolúvel, pelo uso de um reagente multifuncional através de ligações cruzadas 10,11 , por confinamento em matrizes formadas por géis poliméricos 12,13 , por encapsulação em membranas poliméricas 12,14 e em micelas reversas 15,16 .Outra técnica também utilizada, apesar de não manter uma barreira física entre a enzima e o meio reacional, é o sistema bi ou trifásico. Pesquisas recentes têm demonstrado a utilização de novos materiais para imobilização de enzimas, destacando-se os líquidos iônicos com formação de sistemas trifásicos 17-21 , as nanopartículas de sílica funcionalizada e nanocompósitos de sílica-ouro [22][23][24] .Duas das técnicas de imobilização mais utilizadas são, provavelmente, as de inclusão ou microencapsulação através do confinamento dos biocatalisadores em polímeros insolúveis (formando filmes), ou em microcápsulas. Uma das principais vantagens da utilização destas técnicas é que a enzima não interage quimicamente com o polímero, evitando a des...

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“…The substrate specificity of lipases may be attributed to differences in the geometry and size of their active sites (Pleiss et al, 1998). Both forms of cell-bound lipase (acetone powder and released enzyme) showed preferential specificity for higher alkyl chain-length esters (Figure 1).…”

Section: Hydrolysis Of P-nitrophenyl Estersmentioning

confidence: 99%

Differences in hydrolytic abilities of two crude lipases from Geotrichum candidum 4013

Brabcová

Zarevúcká

Macková

2010

Yeast

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The fungus Geotrichum candidum 4013 produces two types of lipases (extracellular and cell-bound). Both enzymes were tested for their hydrolytic ability to pnitrophenyl esters and compounds having a structure similar to the original substrate (triacylglycerols). Higher lipolytic activity of extracellular lipase was observed when triacylglycerols of medium-(C12) and long-(C18) chain fatty acids were used as substrates. Cell-bound lipase preferentially hydrolysed trimyristate (C14). The differences in the abilities of these two enzymes to hydrolyse p-nitrophenyl esters were observed as well. The order of extracellular lipase hydrolysis relation velocity was as follows: p-nitrophenyl decanoate > p-nitrophenyl caprylate > p-nitrophenyl laurate > p-nitrophenyl palmitate > p-nitrophenyl stearate. The cell-bound lipase indicates preference for p-nitrophenyl palmitate. The most striking differences in the ratios between the activity of both lipases (extracellular : cell-bound) towards different fatty acid methyl esters were 2.2 towards methyl hexanoate and 0.46 towards methyl stearate (C18). The Michaelis constant (K m ) and maximum reaction rate (V max ) for p-nitrophenyl palmitate hydrolysis of cell-bound lipase were significantly higher (K m 2.462 mM and V max 0.210 U/g/min) than those of extracellular lipase (K m 0.406 mM and V max 0.006 U/g/min).

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Anatomy of lipase binding sites: the scissile fatty acid binding site

Cited by 433 publications

References 40 publications

Enzymatic synthesis of glyceride esters in solvent-free system: influence of the molar ratio, lipase source and functional activating agent of the support

Enzymatic synthesis of glyceride esters in solvent-free system: influence of the molar ratio, lipase source and functional activating agent of the support

Imobilização de lipases em filme de caseinato de sódio/glicerol: aplicação na síntese de ésteres

Differences in hydrolytic abilities of two crude lipases from Geotrichum candidum 4013

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