Direct micellar systems as a tool to improve the efficiency of aromatic substrate conversion for fine chemicals production.
Whole-cell bioconversion of naphthalene to (+)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene by Escherichia coli JM109(pPS1778) recombinant strain, carrying naphthalene dioxygenase and regulatory genes cloned from Pseudomonas fluorescens N3, in direct micellar systems is optimized as an example of fine chemicals bioproduction from scarcely water-soluble substrates. The oxygen insertion into the aromatic substrate, which stops at the enantiomerically pure cis dihydroxylated product, is performed in direct microemulsion systems, where a non-ionic surfactant stabilizes naphthalene containing oil droplets in an aqueous medium. These media provide an increased substrate solubility so that a homogeneous reaction can be carried out, while not affecting bacteria viability and performances. The influence of the chemical nature of the oil is investigated. The phase behavior of the direct microemulsion system was monitored for three different oils as a function their volume fraction and characterized through light scattering. The addition of isopropyl palmitate, oleic acid, or glyceryl trioleate, 0.6-1.2% v/v to the micellar systems, led to an increase of the substrate concentration in the solution and particularly its bioavailability, allowing faster catalytic conversions. All these systems resulted in being suitable for catalytic conversions of aromatic compounds. Although the nature of the oil does have a deep effect on the phase behavior of the micellar systems, in the present investigation no differences in the yields and in the rates of product formation of the enzymatic system were observed on changing the oil, thus showing that in this case the substrate concentration or bioavailability is not the rate-limiting step.
Berti
D
D
Dipartimento di Chimica, Università di Firenze and CSGI, Florence, Italy.
Randazzo
D
D
Briganti
F
F
Baglioni
P
P
Scozzafava
A
A
Di Gennaro
P
P
Galli
E
E
Bestetti
G
G
D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular / Q000379:methods; D049308:Dioxygenases; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D007700:Kinetics; D008823:Micelles; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D009281:Naphthalenes / Q000737:chemistry* / Q000378:metabolism*; D009284:Naphthols / Q000138:chemical synthesis*; D010105:Oxygenases / Q000235:genetics / Q000378:metabolism*; D011551:Pseudomonas fluorescens / Q000201:enzymology / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism
D008823:Micelles; D009097:Multienzyme Complexes; D009281:Naphthalenes; D009284:Naphthols; D011994:Recombinant Proteins; C031721:naphthalene; C042334:1,2-dihydroxy-1,2-dihydronaphthalene; D010105:Oxygenases; D049308:Dioxygenases; C038703:naphthalene dioxygenase