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1.
X.-J.?Yu Y.-F.?Shen J.?Dong T.?Li C.?Wang Y.-J.?Zhang L.-F.?Wang Y.-C.?Meng Y.?Yang H.-J.?Wang C.-H.?Lei S.?Hu
Previously, we have reported the crystal structures of Fab fragment of Infliximab in complex with TNFα. The structurally identified epitope on TNFα revealed the mechanism of TNFα inhibition by partially overlapping with the TNFα-receptor interface and the possibility to optimize the binding affinity. In this study, we launched a screen of a phage display library to isolate novel anti-TNFα antibodies based on the infliximab epitope. To develop novel anti-TNFα antibodies, structural analysis, the phage display antibody isolation, step by step antibody optimization, CDR residues random mutagenesis, and binding affinity characterization were performed. One of the novel antibodies generated on the backbone of infliximab, Inf3D6, has the superior binding affinity to TNFα, thus, demonstrating the potential for structure guided optimization for improvement of existing antibody-based therapeutics. 相似文献
2.
Paul Christakopoulos Dimitris Kekos Basil J. Macris Peter W. Goodenough Mahalingeshwara K. Bhat 《Biotechnology letters》1994,16(6):587-592
Summary Purified -Glucosidase from Fusarium oxysporum catalysed the hydrolysis and transglycosylation reactions in the presence of cellobiose and gentiobiose. The product of the latter reaction was mainly a triose. The time of incubation, pH and substrate concentration for transglycosylation reaction were optimised. Under optimal conditions, the concentration of glucose and triose reached approximately 15–20 % of the initial substrate concentration. These results suggested that -glucosidase from F.oxysporum is an ideal enzyme for the synthesis of triose in reasonable quantities. 相似文献
3.
4.
Lei Shang Niels Peter K. Nielsen Wolfgang Stelte Jonas Dahl Jesper Ahrenfeldt Jens Kai Holm Maria Puig Arnavat Lars S. Bach Ulrik B. Henriksen 《Bioenergy Research》2014,7(1):87-94
Combined torrefaction and pelletization is used to increase the fuel value of biomass by increasing its energy density and improving its handling and combustion properties. In the present study, a single-pellet press tool was used to screen for the effects of pellet die temperature, moisture content, additive addition, and the degree of torrefaction on the pelletizing properties and pellet quality, i.e., density, static friction, and pellet strength. Results were compared with pellet production using a bench-scale pelletizer. The results indicate that friction is the key factor when scaling up from single-pellet press to bench-scale pelletizer. Tuning moisture content or increasing the die temperature did not ease the pellet production of torrefied wood chips significantly. The addition of rapeseed oil as a lubricant reduced the static friction by half and stabilized pellet production; however, the pellet quality, strength, and density were negatively affected. The pellets produced from pine wood torrefied at 250 and 280 °C were shorter than pellets produced from untreated wood and their quality did not match conventional wood pellet standards. However, the heating value of the torrefied pellets was higher and the particle size distribution after grinding the pellets was more uniform compared to conventional wood pellets. 相似文献
5.
The effect of succinate, acetate, and phosphate on the production of cellobiose dehydrogenase (CDH), cellobiose: quinone oxidoreductase (CBQase), -glucosidase, and protease by Phanerochaete chrysosporium in media containing cotton linters, filterpaper, microcrystalline cellulose, or acid-treated cellulose was investigated. The succinate medium,with an initial pH of 4.5 and with cotton linters as the cellulose source, has been demonstrated to yield the highest levels of CDH (141 U/l) and -glucosidase (237 U/l), and the lowest levels of CBQase (53 U/l). The optimized culture conditions identified here permit isolation of milligram quantities of CDH and -glucosidase from P. chrysosporium. 相似文献
6.
Francislene A. Hasmann Adalberto Pessoa Jr Inês C. Roberto 《Biotechnology Techniques》1999,13(4):239-242
A mathematical model was found to represent the enzyme yield (Y) as a function of pH (X1) and temperature (X2): Y=36.89+10.83X1–12.17X2–6X1X2–17.24 X12–13.24X22. The optimum values for pH and temperature to attain the maximum b-xylosidase recovery (42%) were 3.3 and 24 °C, respectively. 相似文献
7.
Summary Replacement of the regulatory sequence of theBacillus amyloliquefaciens α-amylase gene (AMY1) by the yeast alcohol dehydrogenase gene promoter (ADC1
p) resulted in increased levels of extracellular α-amylase production inSaccharomyces cerevisiae. Negative regulation of glucoamylase synthesis by theSTA10-encoded repressor was alleviated by replacing the nativeSTA2 gene promoter fromS. cerevisiae var.diastaticus withADC1
p. Enhanced degradation of starch was achieved when the modified versions of theAMY1 andSTA2 genes were introduced jointly intoS. cerevisiae. 相似文献
8.
Kumar A Tiwari S Thavaselvam D Sathyaseelan K Prakash A Barua A Arora S Kameswara Rao M 《Protein expression and purification》2012,83(2):226-232
The high level expression of recombinant proteins in Escherichia coli often leads to the formation of inclusion bodies that contain most of the expressed protein held together by non-covalent forces. The inclusion bodies are usually solubilized using strong denaturing agents like urea and guanidium hydrochloride. In this study recombinant Omp28 (rOmp28) protein of Brucella melitensis was expressed in two different vector systems and further efficient purification of the protein was done by modification in buffers to improve the yield and purity. Different concentrations of Triton X-100 and β-mercaptoethanol were optimized for the solubilization of inclusion bodies. The lysis buffer with 8M urea alone was not sufficient to solubilize the inclusion bodies. It was found that the use of 1% Triton X-100 and 20mM β-mercaptoethanol in lysis and wash buffers used at different purification steps under denaturing conditions increased the yield of purified rOmp28 protein. The final yield of purified protein obtained with modified purification protocol under denaturing conditions was 151 and 90mg/l of the culture or 11.8 and 9.37mg/g of wet weight of cells in pQE30UA and pET28a(+) vector respectively. Thus modified purification protocol yielded more than threefold increase of protein in pQE30UA as compared with purification by conventional methods. 相似文献
9.
Here we present the optimization of PCR conditions for microsatellite analysis of coniferous trees. The use of touchdown protocol
for annealing resulted in a high success rate for optimization using fewer temperature profiles. The use of SYBR¢ Green gel stain to detect PCR products in agarose gels was more sensitive than ethidium bromide. This is valuable for determining
the success of PCR reactions and estimating the amount of PCR products formed—which is crucial in determining the dilution
required to produce bands of similar intensity upon silver staining of the polyacrylamide gels. The use of SYBR¢ Gold for staining polyacrylamide gels was not satisfactory in terms of the image quality produced. However, it was comparable
to silver staining in terms of sensitivity, and could possibly be used in cases where the products are present as sharp single
bands. In those cases, the use of SYBR¢ Gold gel stain would save time and money for staining polyacrylamide gels. 相似文献
10.
Noura El-Ahmady El-Naggar S. A. Haroun Eman A. Oweis A. A. Sherief 《Preparative biochemistry & biotechnology》2013,43(7):712-729
Fungi able to degrade agriculture wastes were isolated from different soil samples, rice straw, and compost; these isolates were screened for their ability to produce β-glucosidase. The most active fungal isolate was identified as Talaromyces pinophilus strain EMOO 13-3. The Plackett–Burman design is used for identifying the significant variables that influence β-glucosidase production under solid-state fermentation. Fifteen variables were examined for their significances on the production of β-glucosidase in 20 experimental runs. Among the variables screened, moisture content, Tween 80, and (NH4)2SO4 had significant effects on β-glucosidase production with confidence levels above 90% (p < 0.1). The optimal levels of these variables were further optimized using Box–Behnken statical design. As a result, the maximal β-glucosidase activity is 3648.519 U g?1, which is achieved at the following fermentation conditions: substrate amount 0.5 (g/250 mL flask), NaNO3 0.5 (%), KH2PO4 0.3 (%), KCl 0.02 (%), MgSO4 · 7H2O 0.01 (%), CaCl2 0.01 (%), yeast extract 0.07 (%), FeSO4 · 7H2O 0.0002 (%), Tween 80 0.02 (%), (NH4)2SO4 0.3 (%), pH 6.5, temperature 25°C, moisture content 1 (mL/g dry substrate), inoculum size 0.5 (mL/g dry substrate), and incubation period 5 days. 相似文献
11.
Toshihiro Hamajima Fumie Takahashi Koji Kato Yukihito Sugano Susumu Yamaki Ayako Moritomo Satoshi Kubo Koji Nakamura Kaoru Yamagami Nozomu Hamakawa Koji Yokoo Hidehiko Fukahori 《Bioorganic & medicinal chemistry》2018,26(14):3917-3924
Chemical optimization of pyrazolopyridine 1, focused on cellular potency, isoform selectivity and microsomal stability, led to the discovery of the potent, selective and orally available PI3Kδ inhibitor 5d. On the basis of its desirable potency, selectivity and pharmacokinetic profiles, 5d was tested in the trinitrophenylated aminoethylcarboxymethyl-Ficoll (TNP-Ficoll)-induced antibody production model, and showed higher antibody inhibition than a 4-fold oral dose of the starting compound 1. These excellent results suggest that 5d is a potential candidate for further studies in the treatment of autoimmune diseases and leukocyte malignancies. 相似文献
12.
Ying-Ying Yang Edit Csapó Yong-Liang Zhang Frederik Sü?mann Sarah L. Stebbings Xuan-Ming Duan Zhen-Sheng Zhao Imre Dékány Matthias F. Kling 《Plasmonics (Norwell, Mass.)》2012,7(1):99-106
We have theoretically studied and optimized the field enhancement and temporal response of single and coupled bimetal Ag/Au
core–shell nanoparticles (NPs) with a diameter of 160 nm and compared the results to pure Ag and Au NPs. Very high-field enhancements
with an amplitude reaching 100 (with respect to the laser field centered at 800 nm) are found at the center of a 2-nm gap
between Ag/Au core–shell dimers. We have explored the excitation of the bimetal core–shell particles by Fourier transform-limited
few-cycle optical pulses and identified conditions for an ultrafast plasmonic decay on the order of the excitation pulse duration.
The high-field enhancement and ultrafast decay makes bimetal core–shell particles interesting candidates for applications
such as the generation of ultrashort extreme ultraviolet radiation pulses via nanoplasmonic field enhancement. Moreover, in
first experimental studies, we synthesized small bimetal Ag/Au core–shell NPs and compared their optical response with pure
Au and Ag NPs and numerical results. 相似文献
13.
A truncated Escherichia coli Novablue γ-glutamyltranspeptidase (EcGGT) gene lacking the first 48-bp coding sequence for part of the signal sequence was amplified by polymerase chain reaction and cloned into expression vector pQE-30 to generate pQE-EcGGT. The maximum production of His6-tagged enzyme by E. coli M15 (pQE-EcGGT) was achieved with 0.1 mM IPTG induction for 12 h at 20 °C. The overexpressed enzyme was purified to homogeneity by nickel-chelate chromatography to a specific transpeptidase activity of 4.25 U/mg protein and a final yield of 83%. The molecular masses of the subunits of the purified enzyme were estimated to be 41 and 21 kDa respectively by SDS-PAGE, indicating EcGGT still undergoes the post-translational cleavage even in the truncation of signal sequence. The optimum temperature and pH for the recombinant enzyme were 40 °C and 9, respectively. The apparent K
m and V
max values for γ-glutamyl-p-nitroanilide as γ-glutamyl donor in the transpeptidation reaction were 37.9 μM and 53.7 × 10−3 mM min−1, respectively. The synthesis of L-theanine was performed in a reaction mixture containing 10 mM
L-Gln, 40 mM ethylamine, and 1.04 U His6-tagged EcGGT/ml, pH 10, and a conversion rate of 45% was obtained. 相似文献
14.
以马铃薯脱毒试管苗茎段为转化受体材料,建立并优化了农杆菌介导的马铃薯遗传转化体系.通过农杆菌介导法将玉米淀粉分支酶基因(Starch branching enzyme b,SBEⅡb)的过表达载体转化马铃薯,接种762个茎段,共获得35株抗性植株.经PCR检测获得了4株转基因阳性植株;对转基因植株进一步进行GUS活性组织化学染色,发现转基因植株的茎段与试管薯均被染上蓝色,表明外源SBEⅡb基因已整合到马铃薯基因组,且正常表达. 相似文献
15.
Biophysics - It was found that the activity of the NMDA receptors (bursts of action potentials) was blocked after long-term cryopreservation of brain slices at –10°С. To reactivate... 相似文献
16.
Wei Li Yonghan Hu Jianchang Li Jennifer R. Thomason Dianne DeVincentis Xuemei Du Junjun Wu Rajeev Hotchandani Thomas S. Rush Jerauld S. Skotnicki Steve Tam Priya S. Chockalingam Elisabeth A. Morris Jeremy I. Levin 《Bioorganic & medicinal chemistry letters》2009,19(16):4546-4550
Potent 3,4-disubstituted benzofuran P1′ MMP-13 inhibitors have been prepared. Selectivity over MMP-2 was achieved through a substituent at the C4 position of the benzofuran P1′ moiety of the molecule. By replacing a backbone benzene with a pyridine and valine with threonine, compounds (e.g., 44) with greatly reduced plasma protein binding were also obtained. 相似文献
17.
Shweta V. Pawar 《Preparative biochemistry & biotechnology》2018,48(1):24-33
This study explores a novel concept of coproduction of uricase and alkaline protease by Bacillus licheniformis using single substrate in single step. Seven local bacterial strains were screened for uricase production, amongst which B. licheniformis is found to produce highest uricase along with alkaline protease. Optimization of various factors influencing maximum enzyme coproduction by B. licheniformis is performed. Maximum enzyme productivity of 0.386?U/mL uricase and 0.507?U/mL alkaline protease is obtained at 8?hr of incubation period, 1% (v/v) inoculum, and at 0.2% (w/v) uric acid when the organism is cultivated at 25°C, 180?rpm, in a media containing xylose as a carbon source, urea as a nitrogen source, and initial pH of 9.5. The statistical experimental design method of Box–Behnken was further applied to obtain optimal concentration of significant parameters such as pH (9.5), uric acid concentration (0.1%), and urea concentration (0.05%). The maximum uricase and alkaline protease production by B. licheniformis using Box–Behnken design was 0.616 and 0.582?U/mL, respectively, with 1.6- and 1.13-fold increase as compared to one factor at a time optimized media. This study will be useful to develop an economic, commercially viable, and scalable process for simultaneous production of uricase and protease enzymes. 相似文献
18.
Jin Wook Tak Biki Gupta Raj Kumar Thapa Kyu Bong Woo Sung Yub Kim Toe Gyeong Go Yongjoo Choi Ju Yeon Choi Jee-Heon Jeong Han-Gon Choi Chul Soon Yong Jong Oh Kim 《AAPS PharmSciTech》2017,18(4):1125-1134
The aim of our current study was to characterize and optimize loxoprofen immediate release (IR)/sustained release (SR) tablet utilizing a three-factor, three-level Box–Behnken design (BBD) combined with a desirability function. The independent factors included ratio of drug in the IR layer to total drug (X 1), ratio of HPMC to drug in the SR layer (X 2), and ratio of Eudragit RL PO to drug in the SR layer (X 3). The dependent variables assessed were % drug released in distilled water at 30 min (Y 1), % drug released in pH 1.2 at 2 h (Y 2), and % drug released in pH 6.8 at 12 h (Y 3). The responses were fitted to suitable models and statistical validation was performed using analysis of variance. In addition, response surface graphs and contour plots were constructed to determine the effects of different factor level combinations on the responses. The optimized loxoprofen IR/SR tablets were successfully prepared with the determined amounts of ingredients that showed close agreement in the predicted and experimental values of tablet characterization and drug dissolution profile. Therefore, BBD can be utilized for successful optimization of loxoprofen IR/SR tablet, which can be regarded as a suitable substitute for the current marketed formulations. 相似文献
19.
Abstract Random Amplified Polymorphic DNA polymerase chain reaction (RAPD‐PCR) is a fast and easy way of identifying DNA polymorphisms generated from several regions of the genome. This could expedite the process of identifying informative polymorphic markers that may be linked to important genes controlling economic traits. In cattle, failure to obtain consistent amplification patterns in RAPD‐PCR has been a cause for concern. This has been attributed to the fact that decamer primers that are used in RAPD‐PCR reactions are likely to amplify regions of DNA where the primer‐template base pairing has some degree of mismatch and that these mismatches fail to repeat from reaction to reaction. This paper describes the use of tricine buffer along with changes in reaction components and thermal cycling conditions that has yielded consistent and reproducible RAPD‐PCR amplifications using single primers and double primer combinations on bovine DNA. 相似文献
20.
The purpose of this research was to study whether the bioavailability of ursodeoxycholic acid could be improved by administering
ursodeoxycholic acid–phospholipid complex (UDCA–PLC) orally to rats. A central composite design approach was used for process
optimization in order to obtain the acceptable UDCA–PLC. The physicochemical properties of the complex obtained by optimal
parameters were investigated by means of scanning electron microscopy and X-ray diffraction. The pharmacokinetic parameters
and bioavailability studies were conducted in rats of UDCA after oral administration of UDCA–PLC and UDCA tablet. Multiple
linear regression analysis for process optimization revealed that the acceptable UDCA–PLC was obtained wherein the optimal
values of X
1, X
2 and X
3 were 3, 60°C and 3 h, respectively. The XRD studies of UDCA–PLC obtained by the optimal parameters demonstrated that UDCA
and phospholipids in the UDCA–PLC were combined by non-covalent bonds, not form new compounds. But pharmacokinetic parameters
of the complex in rats were T
max 1.6 h, C
max 0.1346 μg/ml, 11.437 μg·h/ml, respectively. The relative bioavailability of UDCA of UDCA–PLC was increased by 241%,compared with the reference
ursodeoxycholic acid tablet. 相似文献