Mechanism of protein extraction from the solid state by water-in-oil microemulsions

Hayes, Douglas G.

doi:10.1002/(sici)1097-0290(19970320)53:6<583::aid-bit6>3.0.co;2-i

Cited by 29 publications

(12 citation statements)

References 44 publications

(94 reference statements)

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“…Thus, in protein solubilization into a reversed micellar solution by liquid−liquid extraction a favorable result is generally obtained in a lower pH range than pI, when using an anionic surfactant like AOT. Recently, Hayes (1997) has reported the solubilization mechanism of a solid protein by AOT reversed micelles. The mechanism involves three steps: (1) adsorption of reversed micelles to the solid proteins, (2) transfer of the resulting AOT−protein complex from the solid phase to the reversed micellar phase, and (3) formation of protein‐containing reversed micelles accompanied with the exchange reaction between the protein‐containing and the empty reversed micelles.…”

Section: Resultsmentioning

confidence: 99%

Important Parameters Affecting Efficiency of Protein Refolding by Reversed Micelles

et al. 2000

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Refolding of denatured RNase A as a model of inclusion bodies was performed by reversed micelles formulated with sodium di-2-ethylhexyl sulfosuccinate (AOT) in isooctane. In the novel refolding process, a solid-liquid extraction was utilized as an alternative to the ordinary protein extraction by reversed micelles based on a liquid-liquid extraction. First, the effects of operational parameters such as concentration of AOT, W(o) (= [H(2)O]/[AOT]), and pH were examined on the solubilization of solid denatured proteins into a reversed micellar solution. The solubilization was facilitated by a high AOT concentration, a high W(o) value, and a high pH in water pools. These conditions are favorable for the dispersion of the solid protein aggregates in an organic solvent. Second, the renaturation of the denatured RNase A solubilized into the reversed micellar solution was conducted by addition of glutathione as a redox reagent. A complete renaturation of RNase A was accomplished by adjusting the composition of the redox reagent even at a high protein concentration in which protein aggregation would usually occur in aqueous media. In addition, the renaturation rates were improved by optimizing water content (W(o)) and the pH of water pools in reversed micelles. Finally, the recovery of renatured RNase A from the reversed micellar solution was performed by adding a polar organic solvent such as acetone into the reversed micellar solution. This precipitation method was effective for recovering proteins from reversed micellar media without any significant reduction in enzymatic activity.

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Section: Resultsmentioning

confidence: 99%

Important Parameters Affecting Efficiency of Protein Refolding by Reversed Micelles

et al. 2000

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“…Lysozyme strongly interacts with ionic surfactants (Hayes 1997;Lye et al 1995) while chymotrypsin resides in the interior of microemulsion droplets and to interact much more weakly with surfactants (Hirai et al 1995). The solubility of lysozyme and chymotrypsin were 1.5 and 2 g l -1 , respectively, in 0.15 M surfactant w/omicroemulsion solution that possesses a watersurfactant mole ratio, or w o value, of 11.2, and at 10 g protein l -1 for each in 1.2 M surfactant of w/o-microemulsion solution, w o = 11.2.…”

Section: Resultsmentioning

confidence: 99%

“…On a per surfactant basis, the cyclic ketal surfactant yielded much lower protein solubility than other surfactant systems. For instance, lysozyme and chymotrypsin solubility in 0.05 M Aerosol-OT surfactant solutions (near the maximum allowable concentration for phase disengagement to readily occur) has been reported to be within 8.5-13.3 and 9-13.8 g l -1 , respectively (Hayes 1997;Lye et al 1995;Matzke et al 1992).…”

Section: Resultsmentioning

confidence: 99%

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Solubilization of enzymes in water-in-oil microemulsions and their rapid and efficient release through use of a pH-degradable surfactant

2007

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alpha-Chymotrypsin and lysozyme were solubilized in a water/O-[(2-tridecyl, 2-ethyl-1,3-dioxolan-4-yl)methoxy]-O'-methoxy poly(ethylene glycol) (CK-2,13 surfactant)/isooctane water-in-oil microemulsion solution at 1.5-2 and 10 g l(-1) for 0.15 and 1.2 M: CK-2,13, respectively. Upon contact with an equal volume of 0.1 M: NaH(2)PO(4)/Na(2)HPO(4) buffer, pH 5, a three-phase system (Winsor-III system) was formed, consisting of a surfactant-rich middle phase and aqueous and isooctane-rich "excess" phases. Both enzymes were rapidly released into the aqueous excess phase, with 70% recovery of each in 30 and 60 min for microemulsion solutions containing 0.15 and 1.2 M: surfactant, respectively. The recovered enzymes retained >90% of their original specific activity.

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“…The water content in reverse micelles has been reported to be an important parameter for enzymatic reactions 6, 19, 20. In this paper the water content in reverse micelles is indicated by the molar ratio of water to surfactant in organic phase ( w 0 = [H 2 O]/[Surfactant]).…”

Section: Resultsmentioning

confidence: 99%

Enzymatic esterification of DL‐menthol with propionic acid by lipase from Candida cylindracea

Lü

Chu

Han

et al. 2005

J of Chemical Tech & Biotech

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This work deals with the resolution of DL-menthol with propionic acid by Candida cylindracea lipase (Ccl) in organic solvent reaction systems and a reverse micelles system of sodium 1,4-bis (2-ethylhexyl) sulfosuccinate (AOT). The activity and stability as well as enantioselectivity of the lipase in two systems were studied. The results indicate that the lipase showed higher stability in reverse micelles than in organic solvent, which proved that the reverse micelles system has potential application for maintaining the activity of the enzyme for a long time. This is because lipase molecules can be entrapped in water-containing micro-drops of reverse micelles, avoiding direct-contract with unfavorable organic medium. The enantioselectivity (E > 30, ee p = 92.5) in the two systems is relatively high, although the conversion is moderate. The influence of the characteristic parameters of the two systems, such as pH, temperature, w 0 (molar ratio of water to AOT in reverse micelles systems) and water content (organic solvent) on the conversion of DL-menthol was also investigated.

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Mechanism of protein extraction from the solid state by water-in-oil microemulsions

Cited by 29 publications

References 44 publications

Important Parameters Affecting Efficiency of Protein Refolding by Reversed Micelles

Important Parameters Affecting Efficiency of Protein Refolding by Reversed Micelles

Solubilization of enzymes in water-in-oil microemulsions and their rapid and efficient release through use of a pH-degradable surfactant

Enzymatic esterification of DL‐menthol with propionic acid by lipase from Candida cylindracea

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