Lipase‐catalyzed ester exchange reactions in organic media with controlled humidity

Goderis, H. L.; Ampe, G.; Feyten, M. P.; Fouwé, B. L.; Guffens, W. M.; Cauwenbergh, S. M. Van; Tobback, Paul

doi:10.1002/bit.260300216

Cited by 184 publications

(72 citation statements)

References 30 publications

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“…By using water-miscible organic solvents like glycerol [34], almost 'dry' reverse micelles were created in which solubilized enzymes display a high level of catalytic activity. Considerable activity was recorded also at the lowest water activity tested (a, 0.3) in the lipase-catalyzed hydrolysis of triolein in n-hexane [35] and at a water activity of approximately a, 0.4 in the lipase-catalyzed alcoholysis reaction [36].…”

Section: Discussionmentioning

confidence: 99%

Reverse micelles as a water‐property‐control system to investigate the hydration/activity relationship of α‐chymotrypsin

Dorovska-Taran

Veeger

Visser

1993

European Journal of Biochemistry

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a-Chymotrypsin, solubilized in hydrated reverse micelles of sodium bis(2-ethylhexyl)sulfosuccinate (AOT ) in n-octane, was used as a model system for studying the involvement of different water structures (strongly bound water, disordered water, water clusters and bulk water) in the development of the catalytically active conformation of the enzyme. Results presented in this study indicate a characteristic dependence of the stability/activity profile on the water content of the reverse-micellar system for values of w, of approximately 5 (w, is defined as [H,O]/[AOT]). The results are consistent with heat-capacity measurements for proteins. At very low w, values, the conformation of a-chymotrypsin changes to a very rigid structure in comparison to the structure observed in water. This is demonstrated by the overall center of gravity of the tryptophan fluorescence spectrum of the enzyme at w, = 0.65, which is blue shifted in comparison to the spectrum in bulk water indicating that the enzyme is in an apolar environment. In the absence of a hydration shell, the protein is to a great extent frozen and inactive. A small increase in the level of enzyme hydration (up to w, = 2.3) causes an increase in the amount of strongly bound water associated with the enzyme and the enzyme displays a high catalytic activity. Upon further addition of water, a new unstable water structure with unfavourable enthalpy is developed and the enzyme activity declines, reaching a minimum at w, = 5.1. A new increase of water content within a relatively small range, w, = 5-8, causes a dramatic increase in enzymic activity, reminiscent of a cooperative hydration dependence. In the range w, = 10-29, the effect of hydration on the activity is complete which shows that the enzyme activity depends on the amount of water in contact with the enzyme and not on the total amount of bulk water in the system. The experimental results on enzyme incubation at different w, values followed by dilution to constant high w,, are indicative of inactive conformational substates of a-chymotrypsin. It is demonstrated that highly active enzyme conformations at very low w, values occur via an induced fit mechanism of substrate binding.Protein hydration plays a critical role in the catalytic activity and conformational stability of enzymes [l -31. From the analysis of hydrogen-isotope exchange data in proteins, it appears that the hydrogen-bonding network is a major factor in protein structure and function [4]. From the ability of the highly polar solvent methanol to desorb only 62% of water bound to enzymes [5], it has been suggested that at least two types of enzyme-bound water exist. The first type is structurally bound water. The second type is relatively easily removable water. It was shown that serine proteases contain an unusually large number of internal water molecules that are hydrogen bonded to the protein [6]. The positions of 7 out of 13 internally bound water molecules as found in ychymotrypsin are common sites for bound water in both achymotrypsin, diiso...

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Section: Discussionmentioning

confidence: 99%

Reverse micelles as a water‐property‐control system to investigate the hydration/activity relationship of α‐chymotrypsin

Dorovska-Taran

Veeger

Visser

1993

European Journal of Biochemistry

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“…Microbial lipases constitute an important group of biotechnologically valuable enzymes, mainly because of versatility of their applied properties and ease of mass production (13,28,42). Besides their industrial applications, novel biotechnological applications have been successfully established using lipases for the synthesis of biopolymers and biodiesel, the production of enantiopure pharmaceuticals, agrochemicals and flavor compounds (16).…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of extracellular lipase from a new strain: Pseudomonas aeruginosa SRT 9

Borkar¹,

Bodade²,

Rao³

et al. 2009

Braz. J. Microbiol.

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An extra cellular lipase was isolated and purified from the culture broth of Pseudomonas aeruginosa SRT 9 to apparent homogeneity using ammonium sulfate precipitation followed by chromatographic techniques on phenyl Sepharose CL-4B and Mono Q HR 5/5 column, resulting in a purification factor of 98 fold with specific activity of 12307.8 U/mg. The molecular weight of the purified lipase was estimated by SDS-PAGE to be 29 kDa with isoelectric point of 4.5. Maximum lipase activity was observed in a wide range of temperature and pH values with optimum temperature of 55ºC and pH 6.9. The lipase preferably acted on triacylglycerols of long chain (C14-C16) fatty acids. The lipase was inhibited strongly by EDTA suggesting the enzyme might be metalloprotein. SDS and metal ions such as Hg , Ag 2+ and Fe 2+ decreased the lipase activity remarkedly. Its marked stability and activity in organic solvents suggest that this lipase is highly suitable as a biotechnological tool with a variety of applications including organo synthetic reactions and preparation of enantiomerically pure pharmaceuticals. The Km and Vmax value of the purified enzyme for triolein hydrolysis were calculated to be 1.11 mmol/L and 0.05 mmol/L/min respectively.

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“…Use of Molecular Sieves: The initial water activities of the substrates were adjusted according to Goderis et al 27 by adding the molecular sieves in the reaction mixture. It was found that the esterification reaction was stable when mass equivalent of molecular sieves was used.…”

Section: Resultsmentioning

confidence: 99%

Sucrose Derivatives Preparation using Thermomyces lanuginosus Lipase and Their Application

Ashrafuzzaman¹,

Pyo²,

Cheong³

2014

Bulletin of the Korean Chemical Society

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We immobilized Thermomyces lanuginosus lipase to catalyze transesterification reaction in DMF. This lipase was selected after screening among other commercial lipases. We found that prepared immobilized lipase is particularly useful for preparation of 6-O-acylsucrose with higher conversion rate even in 10 g scale. Several solvents were evaluated for selective transesterification reaction. We noticed that the immobilized lipase retained more than 80 % activity after 5 cycles of 96 h reaction. A general method was also developed to purify the products using simple crystallization and precipitation process. Furthermore, 6-O-vinyladipoylsucrose was subjected to synthesis of the corresponding polymer by radical initiator. The sucrose branched polymer can be used further for evaluation of its biodegradability and other biological applications.

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Lipase‐catalyzed ester exchange reactions in organic media with controlled humidity

Cited by 184 publications

References 30 publications

Reverse micelles as a water‐property‐control system to investigate the hydration/activity relationship of α‐chymotrypsin

Reverse micelles as a water‐property‐control system to investigate the hydration/activity relationship of α‐chymotrypsin

Purification and characterization of extracellular lipase from a new strain: Pseudomonas aeruginosa SRT 9

Sucrose Derivatives Preparation using Thermomyces lanuginosus Lipase and Their Application

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