The price of informality : how informal finance schemes defaulted in China, 1989–2015.

The default of a large number of informal finance schemes in China has caused enormous financial losses, and therefore has potential social and political significance. Analysing 354 defaulted schemes from 1989 to 2015, this study defines how they differ from other types of informal finance. It also produces an ideal-type representation of the default process and concludes that the default results from greed, increasing financial pressure at the individual level and private enterprises’ restricted access to state bank loans at the institutional level. China’s financial system should be more flexible in order to prevent further financial losses through informal financial relations

Optimization of ethylene glycol production from (d)-xylose via a synthetic pathway implemented in Escherichia coli

BACKGROUND: Ethylene glycol (EG) is a bulk chemical that is mainly used as an anti-freezing agent and a raw material in the synthesis of plastics. Production of commercial EG currently exclusively relies on chemical synthesis using fossil resources. Biochemical production of ethylene glycol from renewable resources may be more sustainable. RESULTS: Herein, a synthetic pathway is described that produces EG in Escherichia coli through the action of (d)-xylose isomerase, (d)-xylulose-1-kinase, (d)-xylulose-1-phosphate aldolase, and glycolaldehyde reductase. These reactions were successively catalyzed by the endogenous xylose isomerase (XylA), the heterologously expressed human hexokinase (Khk-C) and aldolase (Aldo-B), and an endogenous glycolaldehyde reductase activity, respectively, which we showed to be encoded by yqhD. The production strain was optimized by deleting the genes encoding for (d)-xylulose-5 kinase (xylB) and glycolaldehyde dehydrogenase (aldA), and by overexpressing the candidate glycolaldehyde reductases YqhD, GldA, and FucO. The strain overproducing FucO was the best EG producer reaching a molar yield of 0.94 in shake flasks, and accumulating 20 g/L EG with a molar yield and productivity of 0.91 and 0.37 g/(L.h), respectively, in a controlled bioreactor under aerobic conditions. CONCLUSIONS: We have demonstrated the feasibility to produce EG from (d)-xylose via a synthetic pathway in E. coli at approximately 90 % of the theoretical yield. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12934-015-0312-7) contains supplementary material, which is available to authorized users

A generic HTS assay for kinase screening: Validation for the isolation of an engineered malate kinase

<div><p>An end-point ADP/NAD⁺ acid/alkali assay procedure, directly applicable to library screening of any type of ATP-utilising/ADP producing enzyme activity, was implemented. Typically, ADP production is coupled to NAD⁺ co-enzyme formation by the conventional addition of pyruvate kinase and lactate dehydrogenase. Transformation of enzymatically generated NAD⁺ into a photometrically active alkali derivative product is then achieved through the successive application of acidic/alkali treatment steps. The assay was successfully miniaturized to search for malate kinase activity in a structurally-guided library of LysC aspartate kinase variants comprising 6,700 clones. The screening procedure enabled the isolation of nine positive variants showing novel kinase activity on (L)-malate, the best mutant, LysC V115A:E119S:E434V exhibited strong substrate selectivity for (L)-malate compared to (L)-aspartate with a (<i>k</i>_cat/<i>K</i>_m)_malate/(<i>k</i>_cat/<i>K</i>_m)_aspartate ratio of 86. Double mutants V115A:E119S, V115A:E119C and E119S:E434V were constructed to further probe the origins of stabilising substrate binding energy gains for (L)-malate due to mutation. The introduction of less sterically hindering side-chains in engineered enzymes carrying E119S and V115A mutations increases the effective volume available for substrate binding in the catalytic pocket. Improved binding of the (L)-malate substrate may be assisted by less hindered movement of the Phe184 aromatic side-chain. Additional favourable long-range electostatic effects on binding arising from the E434V surface mutation are conditionally dependent upon the presence of the V115A mutation close to Phe184 in the active-site.</p></div

Additional file 1. of Optimization of ethylene glycol production from (d)-xylose via a synthetic pathway implemented in Escherichia coli

Log2 transformed expression levels of candidate glycolaldehyde reductase

Principle of the end-point alkali assay method for the high-throughput screening of kinase activity.

<p>Principle of the end-point alkali assay method for the high-throughput screening of kinase activity.</p

Screening assay set-up.

<p>(A) Absorbance spectra of NADH and NAD⁺ co-enzyme solutions, at an initial concentration of 0.5 mM, during the sequential acid/alkali treatment. UV-visible spectra were recorded on solutions of the pure co-enzyme forms (blue curves), after acid (red curves) and alkali treatment (green curves). (B) Absorbance at 360 nm, determined in microplate format, as a function of NAD⁺ alkali derivative concentration assuming the total conversion of the NAD⁺ co-enzyme initially present. (C) NAD⁺ alkali derivative calibration curve, determined in microplate format, using solutions mimicking a reaction medium, typically containing inactivated enzyme crude extract, aspartate, ATP and a varying NADH/NAD⁺ co-enzyme ratio at a fixed final total concentration of the reduced and oxidised forms of 1.5 mM.</p

Calculated (L)-malate electrostatic binding interaction energies in the V115A:E119S and V115A:E119S:E434V Ec-LysC mutant ternary complexes with Mg-ADP.

<p>Calculated (L)-malate electrostatic binding interaction energies in the V115A:E119S and V115A:E119S:E434V Ec-LysC mutant ternary complexes with Mg-ADP.</p

Utilisation of additional (L)-malate binding energy gained by LysC mutation in stabilising enzyme-substrate and enzyme transition-state complexes.

<p>Utilisation of additional (L)-malate binding energy gained by LysC mutation in stabilising enzyme-substrate and enzyme transition-state complexes.</p