ABSTRACT
Fatty acid synthase (FAS) catalyses the reaction between acetyl-CoA and malonyl-CoA to produce fatty acids. It is one of the most important enzyme in lipid biosynthesis. FAS of the oleaginous yeast Rhodosporidium toruloides has two acyl carrier protein (ACP) domains and a distinct subunit composition compared with FASs of other species. As ACP is a protein cofactor crucial for fatty acid chain elongation, more ACPs in the FAS may facilitate the reaction. To study the biochemical and structural properties of this novel FAS from R. toruloides, plasmids were constructed and transformed into Escherichia coli BL21 (DE3). The strain ZWE06 harboring plasmids pET22b-FAS1 and pET24b-FAS2 could co-overexpress the two subunits. The recombinant FAS was purified by sequentially using ammonium sulphate precipitation, sucrose density gradient centrifugation and anion exchange chromatography. The specific activity of the recombinant FAS was 548 mU/mg. The purified complex would be used to study enzyme kinetics and protein structure of FAS, and heterogeneous expression and purification will facilitate revealing the mechanism of this novel FAS with double ACPs.
Subject(s)
Acyl Carrier Protein , Basidiomycota , Chromatography , Escherichia coli , Metabolism , Fatty Acid Synthases , Genetics , Fatty Acids , Plasmids , Recombinant Proteins , GeneticsABSTRACT
Objective To evaluate the efficacy and safety of CT-guided percutaneous celiac plexus block (NCPB) using 25 G controllable curved needle together with 22 G straight needle in treating refractory carcinomatous upper abdominal pain. Methods A total of 18 patients with advanced refractory carcinomatous upper abdominal pain were enrolled in this study. The carcinomatous upper abdominal pain failed to the three-step analgesic therapy. Guided by CT scan, percutaneous injection of ethanol with a 25 G controllable curved needle to destroy celiac plexus was carried out in all patients. According to WHO pain relief standards, the relieving degree of pain was evaluated before NCPB and 2 weeks, one, 2, 3 and 6 months after NCPB. The results were analyzed. Results The technical success rate was 100%. The short-term (within 2 weeks) efficacy rate was 88.8%and the complete remission rate was 38.8%. The long-term (over 3 months) efficacy rate was 50% and the complete remission rate was 20%. No severe complications occurred. Conclusion For refractory carcinomatous upper abdominal pain, CT-guided percutaneous celiac plexus block is a simple, safe and effective treatment.
ABSTRACT
Sclareol is a member of labdane type diterpenes mostly used as fragrance ingredient. To enable microbial production of sclareol, synthetic pathways were constructed by incorporating labdenediol diphosphate synthase (LPPS) and terpene synthase (TPS) of the plant Salvia sclarea into Saccharomyces cerevisiae. It was found that sclareol production could be benefited by overexpression of key enzyme for precursor biosynthesis, construction of fusion protein for substrate channeling, and removal of signal peptides from LPPS and TPS. Under optimal shake flask culture conditions, strain S6 produced 8.96 mg/L sclareol. These results provided useful information for development of heterologous hosts for production of terpenoids.