• Journal of Microbiology
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• v.39 no.4
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• pp.305-313
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• 2001

The spr D gene encoding Strptomyces griseus protease D(SGPD); a chymotrypsin-like proteae, was cloned from Strptomyces griseus IFO13350 and sequence. Most of the amino-acid sequence deduced from the nucleotide sequence is idential to that Strptomyces griseus IMRU3499 except that one amino acid has been deleted and Trp 369 has been substituted into Cys369 in the SGPD from S. griseus IFO13350 without affecting the protease activity. The spr D gene was overexpressed in Streptomyce liv-idans TK24 as a heterologous host. Various media with different compositions were also used to max-imize the productivity of SGPD inthe heterologous host. The SGPD productivity was best when the transformant S. lividans TK24 was cultivated in R2YE medium. The relative chymotrypsin activity of the culture broth measured with an artificial chromogenic substrate, N-scuccinyl-ala-ala-pro-phe-p-nitroanilide, was 16 units/ml. A high level of SGPD was also produced in YEME and SAAM medial but it was relatively lower that in R2YE medium and negligible amounts of SGPD were produced in GYE, GAE and Benedict media. The growth of S. lividans reacted the maximum level of cell mass at days 3 and 4 of the culture, but SGPD production started in the stationary phase of cell growth and kept increase in till the 10$^{th}$ day of culture in R2YE and YEME medium, but in GYE media the productivity reached maximum level at 8days of cultivation. The introduction of the spr D gene into S. lividans TK24 triggered biosyntheis of the pigmented antibiotic , actinorhodin, which implies some protease may paly a very improtant role in secondary-metabolite formation in sStreptomyces.

• Journal of Biosystems Engineering
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• v.42 no.4
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• pp.251-257
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• 2017

Purpose: A chisel mounted on working implement, such as agricultural machinery used in irregular farming conditions, is subjected to highly variable fatigue loading during work. To ensure the safety of the chisel on a working implement for the duration of its service life, fatigue testing must be performed with the proper fatigue test load conditions. In this study, working loads for a chisel were developed by reconstructing loads from strain gage data collected during field tests and used to conduct fatigue tests on the chisel component. Methods: FE analysis with nCode software was utilized to select the proper quantity and locations of strain gages for load measurements. A fatigue test was performed to experimentally verify the fatigue strength of the chisel and to evaluate the validity of the load history developed with the load reconstruction technique. Results: A strain history for the chisel was obtained from data collected during field tests. The data was filtered for the 14-16 km/h speed range, connected, and merged. The chisel load history was developed using the load reconstruction technique. The resulting load history was expressed as a load spectrum using the rain-flow counting method. Conclusions: A fatigue test was conducted on a chisel under a constant load condition with an equivalent load amplitude and number of cycles, as calculated by Miner's Rule for linear damage accumulation. During the fatigue test, there were no cracks at any position. It is concluded that the fatigue test method proposed in this study can be utilized successfully as a durability evaluation method for the chisel.

• Journal of Pharmaceutical Investigation
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• v.30 no.1
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• pp.21-26
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• 2000

Bioavailability of ipriflavone (3-phenyl-7-isopropoxy-4H-I-benzopyran-4-one, IP), an antiosteoporotic drug with poor water-solubility, was studied for various types of pharmaceutical preparation in SD rats. The IP preparation types included (1) intact IP, (2) freezer milled IP (FIP), (3) freezer milled IP physically mixed with freezer milled poly-N-vinylpyrrolidone (PVP) (FIP+FPVP) and (4) spray-dried IP with PVP (SIP). Upon oral administration, SIP showed significantly higher absorption and elimination half-lives and the lag time $(t_{lag})$ than those of FIP+FPVP (approximately 2-fold). These results may be due to a sustained releasing effect of IP in the gastrointestinal tract by spray-drying with PVP. The $C_{max}$ of SIP was about 2 and 10 times higher than those of FIP+FPVP and FIP, respectively. The AUC of SIP was about 6 times higher than that of FIP+FPVP and 60 times that of FIP. Scanning electron microscopy (SEM) showed that SIP consisted of the finest particle size and minimal aggregation than other IP preparations. It is concluded that the IP formula prepared by the spray-drying method with PVP is the most effective approach to the improvement of bioavailability of IP.

• Journal of Crop Science and Biotechnology
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• v.10 no.4
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• pp.201-210
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• 2007

Soybean [Glycine max(L.) Merr.] oil is versatile and used in many products. Modifying the fatty acid profile would make soy oil more functional in food and other products. The ideal oil with the most end uses would have saturates(palmitic + stearic acids) reduced from 15 to < 7%, oleic acid increased from 23 to > 55%, and linolenic acid reduced from 8 to < 3%. Reduced palmitic acid(16:0) is conditioned by three or more recessive alleles at the Fap locus. QTLs for reduced palmitic acid have mapped to linkage groups(LGs) A1, A2, B2, H, J, and L. Genes at the Fad locus control oleic acid content(18:1). Six QTLs($R^2$=4-25%) for increased 18:1 in N00-3350(50 to 60% 18:1) explained four to 25% of the phenotypic variation. M23, a Japanese mutant line with 40 to 50% 18:1 is controlled by a single recessive gene, ol. A candidate gene for FAD2-1A can be used in marker-assisted breeding for high 18:1 from M23. Low linolenic acid(18:3) is desirable in soy oil to reduce hydrogenation and trans-fat accumulation. Three independent recessive genes affecting omega-3 fatty acid desaturase enzyme activity are responsible for the lower 18:3 content in soybeans. Linolenic acid can be reduced from 8 to about 4, 2, and 1% from copies of one, two, or three genes, respectively. Using a candidate gene approach perfect markers for three microsomal omega-3 desaturase genes have been characterized and can readily be used in for marker assisted selection in breeding for low 18:3.

• Journal of the Korean GEO-environmental Society
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• v.14 no.5
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• pp.41-47
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• 2013

Well-graded and dense soils have good engineering properties. Unsaturated soil properties such as shear strength, compressibility and permeability are closely related to the soil-water characteristic curve of the soil. Therefore it is important to study the effects of the coefficient of uniformity and the porosity on the soil-water characteristic curve of the sandy soils, which are also related to the grain size distribution and the density of the soil, respectively. In this study soil-water characteristic curves (SWCCs) for six sandy soil specimens were investigated using Tempe pressure cells. The test data were best-fitted to Fredlund and Xing equation. The obtained fitting parameters and the characteristic points of SWCCs were discussed and correlated with the porosity and the coefficient of uniformity of the specimens. The results show that the smaller the porosity of the specimen becomes, the larger the value of the residual matric suction becomes, whereas the larger the coefficient of uniformity of the specimen becomes, the larger the value of the residual matric suction becomes. Regardless of the coefficient of uniformity, the smaller the porosity of the specimen, the flatter the max. slope of SWCC.

• Journal of Microbiology
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• v.44 no.3
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• pp.363-368
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• 2006

In this study, native Byadyrhizobium strains were isolated from the host plant, Glycine max, harvested from fields in Madhya Pradesh, India, and were typed by Iytic rhizobiophages. Eight indigenous (Soy2, ASR011, ASR031, ASR032, MSR091, ISR050, ISR076 and ISR078) and two exotic strains (USDA123 and CB1809), all of which evidenced a distinct reaction with six phages, were employed in this study. The symbiotic interaction of these strains was studied initially using soybean cultivar JS335 in a sand culture in a controlled environment, and the efficiency was assessed based on the nodule number, nodule dry weight, plant dry weight, nitrogenase activity, and total accumulation of N per plant. Symbiotic effectiveness was found to be highest with the native phage-sensitive isolate ASR011, whereas it was at a minimum with the phage-resistant isolates, ISR050 and ISR078. Additionally, the effectiveness of these strains was evaluated using six soybean cultivars belonging to different maturity groups; namely, Brags, Lee, Pusa20, PK416, JS33S and NRC37. Analysis of variance data evidenced significant differences due to both symbionts, for the majority of the tested parameters. The CB1809, USDA123, and ASR011 strains evidenced relatively superior symbiotic effectiveness with soybean cultivars Brags, Lee and JS335. Strain ISR078 evidenced no significant responses with any of the cultivars. The ASR031 strain performed moderately well with all tested cultivars. The symbiotic response of all the strains was quite poor with cultivar PK416. Our studies showed that a significant relationship existed between the phage sensitivity and symbiotic efficiency of the bacterial strains with the host-cultivars.

• The Korean Journal of Physiology and Pharmacology
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• v.11 no.3
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• pp.121-127
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• 2007

In this study, we investigated the therapeutic effects of a novel formulation of low-dose calcium and vitamin $D_3$ blended with Rehmannia glutinosa Libosch and Eleutherococcus senticosus Max (RE+), in postmenopausal women. The controls were given either a placebo or high dose calcium and vitamin $D_3$ (Ca + D). Bone mineral density (BMD) in the L2-3 lumber spines and femur regions was assessed, and serum osteocalcin, bone-specific alkaline phosphatase (BALP), and cross-linked N-telopeptide of type I collagen (NTx) were used as markers of osteoblast and osteoclast activity. Furthermore, all variables were measured before and after 6 and 12 months of treatment. The osteocalcin level was higher in the RE+ group, and BALP was almost the same in all groups. Serum NTx was significantly decreased in the RE+ group after 12 months (p<0.05). The NTx in the Ca + D and placebo groups showed no significant change. The decrease of femur BMD was further demonstrated in the placebo group, but significantly increased in the RE+ group after 6 and 12 months of treatment (p<0.05). There were significant differences in the percent changes of femur BMD between the placebo and RE+ groups (p<0.01) and Ca+D and RE+ groups (p<0.05). The decrease of spine BMD in the placebo group was inhibited both in the Ca + D and RE+ groups, however, there was significant difference only between the placebo and RE+ groups (p<0.05). These findings suggest that continuous oral therapy of the RE+ formulation reduces rapidly decreasing bone mineral density in postmenopausal women more effectively than high doses of calcium and vitamin $D_3$ alone by inhibiting osteoclastic activity. Therefore, it seems that the RE+ has its own antiosteoporotic effects. We suggest larger clinical studies to determine the most efficacious dosage and benefits of this novel treatment.

• Journal of Microbiology and Biotechnology
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• v.21 no.3
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• pp.256-262
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• 2011

A 20.9-kDa metalloprotease was isolated from dried fruiting bodies of the wild basidiomycete mushroom Lepista nuda. The N-terminal amino acid sequence of the protease was seen to be ATFVLTAATNTLFTA, thus displaying no similarity with the sequences of previously reported metalloproteases. The protease was purified using a procedure that entailed ion-exchange chromatography on CM-Cellulose, Q-Sepharose, and Mono S, and FPLC-gel filtration on Superdex 75. The protease functioned at an optimum pH of 7.0 and an optimum temperature of $50^{\circ}C$. It was also noted that the protease demonstrated a proteolytic activity of 1,756 U/mg toward casein. The $K_m$ of the purified protease toward casein was 6.36 mg/ml at a pH of 7.0 and with a temperature of $37^{\circ}C$, whereas the $V_{max}$ was 9.11 ${\mu}g\;ml^{-1}\;min^{-1}$. The activity of the protease was adversely affected by EDTA-2Na, suggesting that it is a metalloprotease. PMSF, EGTA, aprotinin, and leupeptin exerted no striking inhibitory effect. The activity of the protease was enhanced by $Fe^{2+}$, but was curtailed by $Cd^{2+}$, $Cu^{2+}$, $Hg^{2+}$, $Pb^{2+}$, $Zn^{2+}$, and $Fe^{2+}$ ions. The protease also exhibited inhibitory activity against HIV-1 reverse transcriptase with an $IC_{50}$ value of 4.00 ${\mu}M$. The $IC_{50}$ values toward hepatoma Hep G2 and leukemia L1210 cells in vitro were 4.99 ${\mu}M$ and 3.67 ${\mu}M$, respectively.

• Journal of Microbiology and Biotechnology
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• v.22 no.5
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• pp.690-698
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• 2012

Cr-B2, a Gram-negative hexavalent chromium [Cr(VI)] reducing bacteria, was isolated from the aerator water of an activated sludge process in the wastewater treatment facility of a dye and pigment based chemical industry. Cr-B2 exhibited a resistance for 1,100 mg/l Cr(VI) and, similarly, resistance against other heavy metal ions such as $Ni^{2+}$ (800 mg/l), $Cu^{2+}$ (600 mg/l), $Pb^{2+}$ (1,100 mg/l), $Cd^{2+}$ (350 mg/l), $ZN^{2+}$ (700 mg/l), and $Fe^{3+}$ (1,000 mg/l), and against selected antibiotics. Cr-B2 was observed to efficiently reduce 200 mg/l Cr(VI) completely in both nutrient and LB media, and could convert Cr(VI) to Cr(III) aerobically. Cr(VI) reduction kinetics followed allosteric enzyme kinetics. The $K_m$ values were found to be 43.11 mg/l for nutrient media and 38.05 mg/l for LB media. $V_{max}$ values of 13.17 mg/l/h and 12.53 mg/l/h were obtained for nutrient media and LB media, respectively, and the cooperativity coefficients (n) were found to be 8.47 and 3.49, respectively, indicating positive cooperativity in both cases. SEM analysis showed the formation of wrinkles and depressions in the cells when exposed to 800 mg/l Cr(VI) concentration. The organism was seen to exhibit pleomorphic behavior. Cr-B2 was identified on the basis of morphological, biochemical, and partial 16S rRNA gene sequencing chracterizations and found to be Acinetobacter sp.

• Journal of Microbiology and Biotechnology
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• v.20 no.12
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• pp.1653-1663
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• 2010

The first gene (${\alpha}$-gal1) encoding an extracellular ${\alpha}$-Dgalactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The ${\alpha}$-gal1 gene consisted of an open reading frame of 1,792 base pairs interrupted by six introns that encoded a mature protein of 452 amino acids, including a 24 amino acid secretory signal sequence. The translated protein had highest identity with other fungal ${\alpha}$-galactosidases belonging to glycosyl hydrolase family 27. The ${\alpha}$-gal1 gene was overexpressed as a secretory protein with an N-terminal histidine tag in the methylotrophic yeast Pichia pastoris. Recombinant ${\alpha}$-Gal1 was secreted into the culture medium as a monomeric glycoprotein with a maximal yield of 10.75 mg/l and purified to homogeneity using Hisbinding nickel-agarose affinity chromatography. The purified enzyme was maximally active at $70^{\circ}C$, pH 4.5, and lost no activity over 10 days at $50^{\circ}C$. ${\alpha}$-Gal1 followed Michaelis-Menten kinetics ($V_{max}\;of\;240.3{\mu}M/min/mg,\;K_m\;of\;0.294 mM$) and was inhibited competitively by galactose ($K_m{^{obs}}$ of 0.57 mM, $K_i$ of 2.77 mM). The recombinant T. emersonii ${\alpha}$-galactosidase displayed broad substrate preference, being active on both oligo- and polymeric substrates, yet had strict specificity for the ${\alpha}$-galactosidic linkage. Owing to its substrate preference and noteworthy stability, ${\alpha}$-Gal1 is of particular interest for possible biotechnological applications involving the processing of plant materials.