In this review, we presented the industrial status of biomanufactured polyhydroxyalkanoates (PHA), including poly (3-hydroxybutyrate) (PHB), poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), poly (3-hydroxybutyrate-co-4-hydroxybutyrate) (P3/4HB)), and poly (3-hydroxybutyrate-3-hydroxycaproate) (PHBH). A lot of modification studies, aimed at solving problems of poor thermal stability, narrow processing window and other drawbacks of PHA, are discussed. The properties of PHA can be optimized by using proper modification method, in order to expand its applications.
3-Hydroxybutyric Acid ; Biotechnology ; Hydroxybutyrates ; Polyesters ; Polyhydroxyalkanoates ; chemistry

The present study aimed to unravel the carbon metabolism pathway of Acinetobacter sp. TAC-1, a heterotrophic nitrification-aerobic denitrification (HN-AD) strain that utilizes poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) as a carbon source. Sodium acetate was employed as a control to assess the gene expression of carbon metabolic pathways in the TAC-1 strain. The results of genome sequencing demonstrated that the TAC-1 strain possessed various genes encoding carbon metabolic enzymes, such as gltA, icd, sucAB, acs, and pckA. KEGG pathway database analysis further verified the presence of carbon metabolism pathways, including the glycolytic pathway (EMP), pentose phosphate pathway (PPP), glyoxylate cycle (GAC), and tricarboxylic acid (TCA) cycle in the TAC-1 strain. The differential expression of metabolites derived from distinct carbon sources provided further evidence that the carbon metabolism pathway of TAC-1 utilizing PHBV follows the sequential process of PHBV (via the PPP pathway)→gluconate (via the EMP pathway)→acetyl-CoA (entering the TCA cycle)→CO₂+H₂O (generating electron donors and releasing energy). This study is expected to furnish a theoretical foundation for the advancement and implementation of novel denitrification processes based on HN-AD and solid carbon sources.
3-Hydroxybutyric Acid ; Carbon/metabolism* ; Polyesters ; Hydroxybutyrates ; Metabolic Networks and Pathways

Halohydrin dehalogenase is of great significance for biodegradation of the chlorinated pollutants, and also serves as an important biocatalyst in the synthesis of chiral pharmaceutical intermediates. A putative halohydrin dehalogenase (HheTM) gene from Tistrella mobilis KA081020-065 was cloned and over-expressed in Escherichia coli BL21 (DE3). The recombinant enzyme was purified by Ni-NTA column and characterized. Gel filtration and SDS-PAGE analysis showed that the native form of HheTM was a tetramer. It exhibited the highest activity at 50 degrees C. The nature and pH of the buffer had a great effect on its activity. The enzyme maintained high stability under the alkaline conditions and below 30 degrees C. HheTM catalyzed the transformation of ethyl(S)-4-chloro-3-hydroxybutyrate in the presence of cyanide, to give ethyl (R)-4-cyano-3-hydroxybutyrate, a key intermediate for the synthesis of atorvastatin.
3-Hydroxybutyric Acid ; chemistry ; Bacterial Proteins ; genetics ; metabolism ; Cloning, Molecular ; Escherichia coli ; Hydrolases ; genetics ; metabolism ; Hydroxybutyrates ; chemistry ; Recombinant Proteins ; genetics ; metabolism ; Rhodospirillaceae ; enzymology ; genetics

OBJECTIVES: To find a method to distinguish exogenous gamma-hydroxybutyrate (GHB) from endogenous GHB by establishing ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) based on exosome for quantitative detection of GHB in the rat blood. METHODS: Adult male SD rats were divided into 1 h, 5 h, 10 h administration group and control group. After 1 h, 5 h and 10 h of single precursor of GHB gamma-butyrolactone (GBL) intraperitoneal injection in administration groups, 5 mL blood was collected from the abdominal aorta. Meanwhile, the control group was given a same dose of normal saline, and 5 mL blood was collected at 1 h. Among the 5 mL blood, 0.5 mL was directly detected by HPLC-MS after pretreatment, and exosomes were extracted from the remaining blood by differential centrifugation and detected. RESULTS: The concentration of GHB in the control group was (87.36±33.48) ng/mL, and the concentration with administration at 1 h, 5 h and 10 h was (110 400.00±1 766.35) ng/mL, (1 479.00±687.01) ng/mL and (133.60±12.17) ng/mL, respectively. The results of exosome detection showed that no peak GHB signal was detected in the control group and the 10 h administration group, and the concentrations of GHB at 1 h and 5 h administration groups were (91.47±33.44) ng/mL and (49.43±7.05) ng/mL, respectively. CONCLUSIONS GHB was detected in blood exosome by UPLC-MS, which indicated that exogenous GHB could be detected in plasma exosomes, while endogenous GHB could not be detected, suggesting that this method may be used as a basis to determine whether there is exogenous drug intake.
4-Butyrolactone/chemistry* ; Animals ; Chromatography, Liquid ; Exosomes/chemistry* ; Hydroxybutyrates/chemistry* ; Male ; Rats ; Rats, Sprague-Dawley ; Sodium Oxybate/analysis* ; Tandem Mass Spectrometry/methods*

The technology of synthesis, extraction and modification of biodegradable polyhydroxybutyrate (PHB) is introduced briefly in this article. It is also summarized that the research progress in application of PHB in drug delivery and tissue engineering.
Biocompatible Materials ; chemistry ; Drug Delivery Systems ; Hydroxybutyrates ; chemistry ; Tissue Engineering

Accumulation of poly(3-hydroxybutyrate) [poly(3HB)] by V. natriegens was studied. Results indicated that V. natriegens used glucose, gluconate, fructose and molasses as carbon sources for poly(3HB) synthesis. When molasses was used, up to 28.4% of poly(3HB) to cellular dry weight was accumulated. The accumulation of poly(3HB) followed, was not simultaneously to, the cell growth. Analysis of the PHA polymerase, beta-ketothiolase, and acetoacetyl-CoA reductase showed that the poly(3HB) accumulation was correlated to the increase of their activities in cells. Poly(3HB) accumulation was also related to the de novo fatty acid synthesis, as revealed by the results that cerulenin, a specific inhibitor to the de novo fatty acid synthesis, significantly reduced accumulation of poly(3HB). Based on the results from this study, the synthetic pathway of poly(3HB) was proposed.
Cerulenin ; pharmacology ; Hydroxybutyrates ; metabolism ; Polyesters ; metabolism ; Vibrio ; metabolism

Polyhydroxyalkanoates (PHAs) have obtained much attention in biomaterial fields due to their similar physicochemical properties to those of the petroleum-derived plastics. Poly(3-hydroxybutyrate-co-lactate) [P(3HB-co-LA)] is one member of the PHAs family, and has better toughness and transparency compared to existing polylactic acid (PLA) and poly[(R)-3-hydroxybutyrate] [P(3HB)]. First, we confirmed the one-step biosynthesis of P(LA-co-3HB) with the lactate fraction of 23.8 mol% by introducing P(3HB-co-LA) production module into Escherichia coli MG1655. Then, the lactate fraction was increased to 37.2 mol% in the dld deficient strain WXJ01-03. The genes encoding the thioesterases, ydiI and yciA, were further knocked out, and the lactate fraction in the P(3HB-co-LA) was improved to 42.3 mol% and 41.1 mol% respectively. Strain WXJ03-03 with dld, ydiI and yciA deficient was used for the production of the LA-enriched polymer, and the lactate fraction was improved to 46.1 mol%. Notably, the lactate fraction in P(3HB-co-LA) from xylose was remarkably higher than from glucose, indicating xylose as a potent carbon source for P(3HB-co-LA) production. Therefore, the deficiency of thioesterase may be considered as an effective strategy to improve the lactate fraction in P(3HB-co-LA) in xylose fermentation.
Escherichia coli/genetics* ; Hydroxybutyrates ; Lactic Acid ; Polyesters ; Polyhydroxyalkanoates ; Xylose

2-Hydroxybutyric acid (2-HBA) is an important intermediate for synthesizing biodegradable materials and various medicines. Chemically synthesized racemized 2-HBA requires deracemization to obtain optically pure enantiomers for industrial application. In this study, we designed a cascade biosynthesis system in Escherichia coli BL21 by coexpressing L-threonine deaminase (TD), NAD-dependent L-lactate dehydrogenase (LDH) and formate dehydrogenase (FDH) for production of optically pure (S)-2-HBA from bulk chemical L-threonine (L-Thr). To coordinate the production rate and the consumption rate of the intermediate 2-oxobutyric acid in the multi-enzyme cascade catalytic reactions, we explored promoter engineering to regulate the expression levels of TD and FDH, and developed a recombinant strain P21285FDH-T7V7827 with a tunable system to achieve a coordinated multi-enzyme expression. The recombinant strain P21285FDH-T7V7827 was able to efficiently produce (S)-2-HBA with the highest titer of 143 g/L and a molar yield of 97% achieved within 16 hours. This titer was approximately 1.83 times than that of the highest yield reported to date, showing great potential for industrial application. Our results indicated that constructing a multi-enzyme-coordinated expression system in a single cell significantly contributed to the biosynthesis of hydroxyl acids.
Escherichia coli/genetics* ; Formate Dehydrogenases ; Hydroxybutyrates ; Threonine Dehydratase

Based on the fermentation analysis of Escherichia coli strains and cheap renewable resources suitable for polyhydroxybutyrate (PHB) production, we constructed a ptsG mutant of Escherichia coli DH5alpha. Application of E. coli DH5alpha mutant together with stress-induced system, we could produce PHB efficiently from cheap renewable sugar mixture by the simultaneous consumption of different sugars. Batch fermentation at lab scale (5 liter) showed that E. coli DH5alpha deltaptsG/pQKZ103 produced PHB from sugar mixture up to 84.6% of cell dry weight in 32 hours; meanwhile, the cell dry weight reached 8.24 g/L.
Escherichia coli ; genetics ; metabolism ; Fermentation ; Genetic Vectors ; genetics ; Hydroxybutyrates ; metabolism ; Metabolic Engineering ; methods ; Mutation ; Polyesters ; metabolism

4-Hydroxybutyric acid (4HB) is a psychotropic drug used for polymer synthesis such as poly (4-hydroxybutyric acid) (P4HB) and poly (3-hydroxybutyric acid-co-4-hydroxybutyric acid) (P3HB-co-4HB). 1,4-butanediol (BD) can be converted to 4-hydroxybutyric acid by alcohol dehydrogenase (DhaT) and aldehyde dehydrogenase (AldD). In this study, high efficiency promoters including T7 promoter and P(Re) promoter were cloned to increase expression of dhaT and aldD, and thus accelerate the conversion from BD to 4HB. A. hydrophila 4AK4 (pZQ01), the recombinant strain under the control of T7 promoter, produced 6.00 g/L 4HB from 10 g/L BD with the productivity increased by 43.20%. While A. hydrophila 4AK4 (pZQ04), the strain under the control of T7 promoter, produced 4.87 g/L 4HB from 10 g/L BD, and the productivity was increased by 16.23%. Thus, the gene expression was increased by T7 and P(Re) promoters, leading to an accelerated biosynthesis of 4HB.
Aeromonas hydrophila ; genetics ; metabolism ; Genetic Engineering ; Hydroxybutyrates ; metabolism ; Promoter Regions, Genetic ; genetics ; Recombination, Genetic