Fatty Acid Production by Enhanced Malonyl-CoA Supply in Escherichia coli.

The expression of exogenous genes encoding acetyl-CoA carboxylase (Acc) and pantothenate kinase (CoaA) in Escherichia coli enable highly effective fatty acid production. Acc-only strains grown at 37 °C or 23 °C produced an approximately twofold increase in fatty acid content, and additional expression of CoaA achieved a further twofold accumulation. In the presence of pantothenate, which is the starting material for the CoA biosynthetic pathway, the size of the intracellular CoA pool at 23 °C was comparable to that at 30 °C during cultivation, and more than 500 mg/L of culture containing cellular fatty acids was produced, even at 23 °C. However, the highest yield of cellular fatty acids (1100 mg/L of culture) was produced in cells possessing the gene encoding type I bacterial fatty acid synthase (FasA) along with the acc and coaA, when the transformant was cultivated at 30 °C in M9 minimal salt medium without pantothenate or IPTG. This E. coli transformant contained 141 mg/L of oleic acid attributed to FasA under noninducible conditions. The increased fatty acid content was brought about by a greatly improved specific productivity of 289 mg/g of dry cell weight. Thus, the effectiveness of the foreign acc and coaA in fatty acid production was unambiguously confirmed at culture temperatures of 23 °C to 37 °C. Cofactor engineering in E. coli using the exogenous coaA and acc genes resulted in fatty acid production over 1 g/L of culture and could effectively function at 23 °C.

Enhancement of fatty acid biosynthesis by exogenous acetyl-CoA carboxylase and pantothenate kinase in Escherichia coli.

OBJECTIVES: To establish a technique for efficient fatty acid production through enhancement of coenzyme A (CoA) biosynthesis and malonyl-CoA supply by introducing exogenous pantothenate kinase (coaA) and acetyl-CoA carboxylase (acc) in Escherichia coli. RESULTS: The expression of acc, obtained from Corynebacterium glutamicum, accumulated 2.2-fold more fatty acids in E. coli. The addition of coaA from Pseudomonas putaida or fatty acid synthase (fasA) from C. glutamicum resulted in a 3.1- and 3.6-fold increase in fatty acid synthesis in E. coli cells, which expressed acc and coaA, or acc and fasA, respectively. The transformants, simultaneously possessing all three genes, produced 5.6-fold more fatty acids. The strain possessing acc, coaA, and fasA stored 691 mg/L of fatty acids, primarily as phospholipids, inside the inner membrane after 72-h cultivation. In addition, 19% of the total CoA pool was occupied by malonyl-CoA. CONCLUSIONS: Increased malonyl-CoA significantly contributed to fatty acid production, and the effect was boosted by the expanded total CoA pool. Manipulation of the intracellular CoA species is effective for fatty acid production in E. coli.

Change in anterior chamber depth following combined pars plana vitrectomy, phacoemulsification, and intraocular lens implantation using different types of intraocular lenses.

PURPOSE: To examine whether the type of intraocular lens (IOL) used in combined pars plana vitrectomy, phacoemulsification, and IOL implantation affects the changes in anterior chamber depth over time. METHODS: A retrospective review was carried out on data from 70 eyes of 70 patients who underwent combined vitrectomy and cataract surgery. Vitrectomy using a 23-gauge system was performed on 66 eyes and using a 25-gauge system on four eyes. The implanted IOLs were the HOYA VA-65BB lens in 38 eyes (6.5-mm group) and the ETERNITY X-70 lens in 32 eyes (7-mm group). Anterior chamber depth was measured using a PENTACAM analyzer before surgery and 1 week, 1 month, and 3 months after surgery. RESULTS: In the 7-mm group, no differences were found in anterior chamber depth between eyes with and without fluid-gas exchange at any point of time after surgery. In the 6.5-mm group, eyes undergoing fluid-gas exchange showed an increase in anterior chamber depth between 1 week and 1 month after surgery. In eyes undergoing fluid-gas exchange, anterior chamber depth 1 week after surgery was shallower in the 6.5-mm group than in the 7-mm group. CONCLUSION: Different types of three-piece IOLs showed different degrees of shift due to fluid-gas exchange.

Masseter muscular weakness affects temporomandibular synovitis induced by jaw opening in growing rats.

OBJECTIVE: To evaluate the influence of impaired masseter function during growth on the development of temporomandibular synovitis. MATERIALS AND METHODS: Sixteen 3-week-old male Wistar rats were classified into four groups. The first group served as control; and in the second group, jaw opening was forced for 3 hours when the rats were 9 weeks old. In the third and fourth groups, the masseter muscles were bilaterally resected at 3 weeks of age, and the rats in the fourth group were additionally forced to open their jaw at 9 weeks of age. All rats were sacrificed at 9 weeks. Temporomandibular joint (TMJ) tissue samples were processed for histology, and evaluated for cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expressions by immunohistochemistry to examine the inflammatory changes in the synovial membrane. RESULTS: The control group showed noninflammatory changes. In the jaw-opening group, vascular dilation and weak COX-2 immunoreactivity were induced by jaw opening in the synovium. In the masseter-resection group, the masseter-resected rats exhibited moderate synovial changes while in the resection with opening group, the masseter-resected rats revealed more significant inflammatory changes including synovial hyperplasia, dilated vasculature, fibrin deposits, and intense immunoreactivity for COX-2 and iNOS, all caused by jaw opening. CONCLUSIONS: These results suggest that masseter activity in the growth period is an important factor in the induction of temporomandibular synovitis.