Polyhydroxyalkanoate (PHA) is a family of biodegradable polymers, and incorporation of different monomers can alter its physical properties. To produce the copolymer poly(3-hydroxybutyrate
-co-
...3-hydroxyvalerate) (P(3HB-
co
-3HV)) containing a high level of 3-hydroxyvalerate (3HV) by altering acetyl-CoA pool levels, we overexpressed an acetyl-CoA acetyltransferase (
atoAD
) in an engineered
E. coli
strain, YH090, carrying PHA synthetic genes
bktB, phaB
, and
phaC
. It was found that, with introduction of
atoAD
and with propionate as a co-substrate, 3HV fraction in PHA was increased up to 7.3-fold higher than a strain without
atoAD
expressed
in trans
(67.9 mol%). By the analysis of CoA pool concentrations in vivo and in vitro using HPLC and LC-MS, overexpression of AtoAD was shown to decrease the amount of acetyl-CoA and increase the propionyl-CoA/acetyl-CoA ratio, ultimately resulting in an increased 3HV fraction in PHA. Finally, synthesis of P(3HB-
co
-3HV) containing 57.9 mol% of 3HV was achieved by fed-batch fermentation of YJ101 with propionate.
Purpose
The purpose of this study was to compare in vivo kinematics, range of motion, and functional outcomes in patients that received either a high-flexion cruciate retaining or a high-flexion ...cruciate substituting knee.
Methods
Forty-eight high-flexion cruciate retaining (CR) and 47 high-flexion cruciate substituting (PS) knees were included in this study and followed for an average of 27 months (24–33). Weight-bearing and non-weight-bearing maximal flexions and functional scores were compared between two groups. For kinematics evaluations, amount of posterior femoral roll-back and internal tibial rotation from 0° to maximal flexion using lateral radiographs under weight-bearing conditions were also compared.
Results
Average weight-bearing maximal flexion was 126.3° in the PS group, which was significantly higher than the 115.0° in the CR group. Average functional scores showed no significant difference between the two groups. In terms of kinematics, the average amount of posterior femoral roll-back during full flexion was 9.6 mm in the PS group and 6.1 mm in the CR group, which was a significant difference. However, internal tibial rotation during full flexion was not significantly different in the two groups.
Conclusion
PS high-flexion TKA provided greater weight-bearing maximal flexion and posterior femoral roll-back than CR high-flexion TKA, although no difference in clinical outcomes was observed between the two prosthesis designs.
Level of evidence
I.
L-Theanine, found in green tea leaves has been shown to positively affect immunity and relaxation in humans. There have been many attempts to produce L-theanine through enzymatic synthesis to ...overcome the limitations of traditional methods. Among the many genes coding for enzymes in the L-theanine biosynthesis, glutamylmethylamide synthetase (GMAS) exhibits the greatest possibility of producing large amounts of production. Thus, GMAS from Methylovorus mays No. 9 was overexpressed in several strains including vectors with different copy numbers. BW25113(DE3) cells containing the pET24ma::gmas was selected for strains. The optimal temperature, pH, and metal ion concentration were 50℃, 7, and 5 mM MnCl 2 , respectively. Additionally, ATP was found to be an important factor for producing high concentration of L-theanine so several strains were tested during the reaction for ATP regeneration. Baker’s yeast was found to decrease the demand for ATP most effectively. Addition of potassium phosphate source was demonstrated by producing 4-fold higher L-theanine. To enhance the conversion yield, GMAS was additionally overexpressed in the system. A maximum of 198 mM L-theanine was produced with 16.5 mmol/l/h productivity. The whole-cell reaction involving GMAS has greatest potential for scale-up production of L-theanine.
Rapamycin, produced by the soil bacterium Streptomyces hygroscopicus, has the ability to suppress the immune system and is used as an antifungal, anti-inflammatory, antitumor, and immunosuppressive ...agent. In an attempt to increase the productivity of rapamycin, mutagenesis of wild-type Streptomyces hygroscopicus was performed using ultraviolet radiation, and the medium composition was optimized using glycerol (which is one of the cheapest starting substrates) by applying Plackett-Burman design and response surface methodology. Plackett-Burman design was used to analyze 14 medium constituents: M100 (maltodextrin), glycerol, soybean meal, soytone, yeast extract, (NH4)2SO4, L-lysine, KH2PO4, K2HPO4, NaCl, FeSO4·7H2O, CaCO3, 2-(N-morpholino) ethanesulfonic acid, and the initial pH level. Glycerol, soytone, yeast extract, and CaCO3 were analyzed to evaluate their effect on rapamycin production. The individual and interaction effects of the four selected variables were determined by Box-Behnken design, suggesting CaCO3, soytone, and yeast extract have negative effects, but glycerol was a positive factor to determine rapamycin productivity. Medium optimization using statistical design resulted in a 45% (220.7 ± 5.7 mg/l) increase in rapamycin production for the Streptomyces hygroscopicus mutant, compared with the unoptimized production medium (151.9 ± 22.6 mg/l), and nearly 588% compared with wildtype Streptomyces hygroscopicus (37.5 ± 2.8 mg/l). The change in pH showed that CaCO3 is a critical and negative factor for rapamycin production.
Background: Painful plantar callosities under lesser metatarsal heads are commonly associated with hallux valgus. The purpose of the present study was to evaluate the prognosis of painful plantar ...callosities after hallux valgus correction without lesser metatarsal osteotomy in hallux valgus deformity. Materials and Methods: Between September 2004 and June 2007, 31 patients (40 feet) underwent proximal chevron first metatarsal osteotomy with a distal soft tissue procedure, with preoperatively painful plantar callosities under lesser metatarsal heads. Clinical results were evaluated using a visual analogue scale (VAS), the American Orthopaedic Foot and Ankle Society (AOFAS) hallux-metatarsophalangeal interphalangeal scales, and a modified 70-point clinical scale. Radiographic evaluations included hallux valgus angle and intermetatarsal angle. Results: Thirty-two (80%) of the 40 feet had no pain and callosity and 5 (12.5%) had no pain but residual plantar callosities, and 3 (7.5%) were not improved at final evaluation. The mean VAS and AOFAS scores were improved from 7.8 ± 1.6 to 1.9 ± 1.5 points and from 53.8 ± 14.2 to 92.6 ± 15.3 points, respectively. In terms of the 70-point clinical scale, overall clinical results were good in 34 feet and fair in 6. The mean hallux valgus and intermetatarsal angles were improved from 36.6 ± 6.2 to 12.5 ± 5.9 degrees and from 17.5 ± 3.9 to 8.6 ± 3.5 degrees, respectively. Conclusion: Painful plantar callosities under the lesser metatarsals in patients with hallux valgus deformity can be improved by hallux valgus correction alone without lesser metatarsal osteotomy.
Level of Evidence: IV, Retrospective Case Series
SCO0948 was found to be the single open reading frame annotated to encode an α-mannosidase (AM1) in Streptomyces coelicolor M145. To characterize the protein, we overexpressed SCO0948 in Escherichia ...coli BL21(DE3). Recombinant AM1, with a molecular weight of 110 kDa, exhibited α-mannosidase activity toward 4-nitrophenyl-α-D-mannopyranoside with a K ₘ of 4.61 mM, a V ₘₐₓ of 101.6 mM/min, and a specific activity of 47.96 U/mg. Treatment of ovalbumin, a glycoprotein, with AM1 resulted in partial deglycosylation, as assessed by glycostaining and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The S. coelicolor deletion mutant for SCO0948 failed to produce α-mannosidase activity, confirming AM1 as the only α-mannosidase in S. coelicolor M145. Interestingly, the deletion mutant and a complementation strain produced lower levels of the antibiotics actinorhodin and undecylprodigiosin in glucose minimal media. The results indicate that AM1 as an α-mannosidase influences deglycosylation and antibiotic production in S. coelicolor M145.
A putative ω-aminotransferase gene, cc3143 (aptA), from Caulobacter crescentus was screened by bioinformatical tools and overexpressed in E. coli, and the substrate specificity of the ...ω-aminotransferase was investigated. AptA showed high activity for short-chain β-amino acids. It showed the highest activity for 3-amino-n-butyric acid. It showed higher activity toward aromatic amines than aliphatic amines. The 3D model of the ω-aminotransferase was constructed by homology modeling using a dialkylglycine decarboxylase (PDB ID: 1DGE) as a template. Then, the ω-aminotransferase was rationally redesigned to increase the activity for 3-amino-3-phenylpropionic acid. The mutants N285A and V227G increased the relative activity for 3-amino-3-phenylpropionic acid to 3-amino-n-butyric acid by 11-fold and 3-fold, respectively, over that of wild type.
