The traditional distillation method for recovery of butanol from fermentation broth is an energy-intensive process. Separation of butanol based on adsorption methodology has advantages in terms of biocompatibility and stability, as well as economy, and therefore gains much attention. However, the application of the commercial adsorbents in the integrated acetone-butanol-ethanol (ABE) fermentation process is restricted due to the low recovery (less than 85%) and the weak capability of enrichment in the eluent (3-4 times). In this study, we investigated the sorption properties of butanol onto three kinds of adsorbents with different polarities developed in our laboratory, that is, XD-41, H-511, and KA-I resin. The sorption behaviors of single component and ABE ternary mixtures presented in the fermentation broths on KA-I resin were investigated. KA-I resin had higher affinity for butanol than for acetone, ethanol, glucose, acetic acid, and butyric acid. Multicomponent ABE sorption on KA-I resin was modeled using a single site extended Langmuir isotherm model. In a desorption study, all the adsorbed components were desorbed in one bed volume of methanol, and the recovery of butanol from KA-I resin was 99.7%. The concentration of butanol in the eluent was increased by a factor of 6.13. In addition, KA-I resin was successfully regenerated by two bed volumes of water. Because of its quick sorption, high sorption capacity, low cost, and ease of desorption and regeneration, KA-I resin exhibits good potential for compatibility with future ABE fermentation coupled with in situ recovery product removal techniques.
A new member of the CYP116B subfamily-P450LaMO-was discovered in Labrenzia aggregata by genomic data mining. It was successfully overexpressed in Escherichia coli, purified, and subsequently characterized spectroscopically, and its catalytic properties were assessed. Substrate profiling of the P450LaMO revealed that it was a versatile catalyst, exhibiting hydroxylation and epoxidation activities as well as O-dealkylation and asymmetric sulfoxidation activities. Diverse compounds, including alkylbenzenes, aromatic bicyclic molecules, and terpenoids, were shown to be hydroxylated by P450LaMO. Such diverse catalytic activities are uncommon for the bacterial P450s, and the P450LaMO-mediated stereoselective hydroxylation of inactivated C-H bonds-ubiquitous and relatively unreactive in organic molecules-is particularly unusual. The self-sufficient nature of P450LaMO, coupled with its broad substrate range, highlights it as an ideal template for directed evolution towards various applications.
Regioselective
hydroxylations of aromatic compounds are useful
reactions but often lack appropriate catalysts. Here a group of P450BM3
mutants (R47I/A82F/A328F, R47L/Y51F/F87V/L188P/I401P, R47I/Y51F/F87V,
R47L/Y51F/F87V/L181Q/L188P/I401P, and R47I/F87V/L188P) were developed
as unique catalysts for the p-hydroxylation of m-alkylphenols 1a–e with
high regioselectivity (91–99%) and conversion (95–99%)
to produce the corresponding useful and valuable m-alkylbenzene-1,4-diols 2a–e, respectively.
The mutated hydroxylases were developed by protein engineering of
P450BM3 monooxygenase via site-directed mutagenesis based on designed
mutations to reshape the substrate binding pocket and access channel.
Several engineered P450BM3 mutants showed good catalytic efficiency
(k
cat/K
M of
234–381 mM–1 min–1) for
the p-hydroxylations of m-alkylphenols 1a–e, respectively. Molecular docking
and simulation gave some insights into the structure-based understanding
of the enhanced regioselectivity and activity for the developed P450BM3
mutants, including the shorter distance between heme-oxygen atom and
C4-carbon (p-position) of substrates than the wild-type
enzyme in the catalytic pockets. Preparative biohydroxylations of m-alkylphenols 1a–e were
demonstrated by using E. coli cells coexpressing
individual P450BM3 mutants and glucose dehydrogenase GDH, giving high-yielding
synthesis of useful and valuable m-alkylbenzene-1,4-diols 2a–e.
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