Enzymatic Synthesis of l-threo-ß-Hydroxy-α-Amino Acids via Asymmetric Hydroxylation Using 2-Oxoglutarate-Dependent Hydroxylase from Sulfobacillus thermotolerans Strain Y0017.

ß-Hydroxy-α-amino acids are useful compounds for pharmaceutical development. Enzymatic synthesis of ß-hydroxy-α-amino acids has attracted considerable interest as a selective, sustainable, and environmentally benign process. In this study, we identified a novel amino acid hydroxylase, AEP14369, from Sulfobacillus thermotolerans Y0017, which is included in a previously constructed CAS-like superfamily protein library, to widen the variety of amino acid hydroxylases. The detailed structures determined by nuclear magnetic resonance and X-ray crystallography analysis of the enzymatically produced compounds revealed that AEP14369 catalyzed threo-ß-selective hydroxylation of l-His and l-Gln in a 2-oxoglutarate-dependent manner. Furthermore, the production of l-threo-ß-hydroxy-His and l-threo-ß-hydroxy-Gln was achieved using Escherichia coli expressing the gene encoding AEP14369 as a whole-cell biocatalyst. Under optimized reaction conditions, 137 mM (23.4 g liter-1) l-threo-ß-hydroxy-His and 150 mM l-threo-ß-hydroxy-Gln (24.3 g liter-1) were obtained, indicating that the enzyme is applicable for preparative-scale production. AEP14369, an l-His/l-Gln threo-ß-hydroxylase, increases the availability of 2-oxoglutarate-dependent hydroxylase and opens the way for the practical production of ß-hydroxy-α-amino acids in the future. The amino acids produced in this study would also contribute to the structural diversification of pharmaceuticals that affect important bioactivities. IMPORTANCE Owing to an increasing concern for sustainability, enzymatic approaches for producing industrially useful compounds have attracted considerable attention as a powerful complement to chemical synthesis for environment-friendly synthesis. In this study, we developed a bioproduction method for ß-hydroxy-α-amino acid synthesis using a newly discovered enzyme. AEP14369 from the moderate thermophilic bacterium Sulfobacillus thermotolerans Y0017 catalyzed the hydroxylation of l-His and l-Gln in a regioselective and stereoselective fashion. Furthermore, we biotechnologically synthesized both l-threo-ß-hydroxy-His and l-threo-ß-hydroxy-Gln with a titer of over 20 g liter-1 through whole-cell bioconversion using recombinant Escherichia coli cells. As ß-hydroxy-α-amino acids are important compounds for pharmaceutical development, this achievement would facilitate future sustainable and economical industrial applications.

Use of a seeder reactor to manage crystal growth in the fluidized bed reactor for phosphorus recovery.

The authors have been engaged in the development of a phosphorus recovery system capable of maintaining high recovery efficiencies, with the chemical cost suppressed. This time, they conducted demonstration tests of a fluidized bed magnesium ammonium phosphate reactor provided with a seeder reactor for the supernatant from anaerobic digestion using a pilot experimental plant with a wastewater treatment capacity of 20 m3/d. For the digestion supernatant with a phosphorus concentration of approximately 300 mg/L, the treated water phosphorus concentration was 10 to 25 mg/L, and the phosphorus recovery efficiency was more than 90%. Relative to the chemical cost in the case of magnesium chloride, the chemical cost in the case of magnesium hydroxide is approximately 40%. Thus, with the new system, it was possible to reduce the running cost while maintaining high recovery efficiencies.

Effect of ultrasonic irradiation on nucleation phenomena in a Na2HPO4.12H2O melt being used as a heat storage material.

The effect of ultrasound on nucleation phenomena in the heat storage material Na2HPO4.12H2O was investigated by determining the primary nucleation probability and induction time, and by looking at heat generation phenomena in the initial stage of nucleation. The experimental results show that the primary nucleation probability dramatically increased, and the induction time decreased under the ultrasound irradiation, and in addition, the rate of temperature rise was dependent upon the ultrasonic output. Based on these results and the theoretical relationship between the number of primary nuclei and the heat generation rate, it is proposed that the number of primary nuclei depends upon the ultrasonic output.

Morphological control of calcium carbonate crystallized in reverse micelle system with anionic surfactants SDS and AOT.

The influence of a surfactant over water on the polymorphism and crystal size of calcium carbonate produced by reaction crystallization in microemulsion systems was investigated in a mixing tank reactor. The crystallization was induced by the reaction between two aqueous micelle solutions (Na2CO3-CaCl2) stabilized by anionic surfactants, SDS (sodium dodecyl sulfate) or AOT (sodium bis(2-ethylhexyl) sulfosuccinate). With increasing surfactant ratio to water, the water-in-oil microemulsion was stably developed and the morphology of the calcium carbonate crystallized in the micelles sharply transformed from calcite to vaterite. The influence of SDS on the polymorphism and crystal size of calcium carbonate was much clearer than that of AOT. In addition, with AOT, certain step changes in the morphology and crystal size occurred around a surfactant ratio to water (R=[H2O]/[surfactant]) of 15 due to a two-phase separation of the microemulsion.