• 제목/요약/키워드: Protein engineering

검색결과 2,930건 처리시간 0.03초

Proteomics-driven Identification of Putative AfsR2-target Proteins Stimulating Antibiotic Biosynthesis in Streptomyces lividans

  • Kim Chang-Young;Park Hyun-Joo;Kim Eung-Soo
    • Biotechnology and Bioprocess Engineering:BBE
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    • 제10권3호
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    • pp.248-253
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    • 2005
  • AfsR2, originally identified from Streptomyces lividans, is a global regulatory protein which stimulates antibiotic biosynthesis. Through its stable chromosomal integration, the high level of gene expression of afsR2 significantly induced antibiotic production as well as the sporulation of S. lividans, implying the presence of yet-uncharacterized AfsR2-target proteins. To identify and evaluate the putative AfsR2-target proteins involved in antibiotic regulation, the proteomics-driven approach was applied to the wild-type S. lividans and the afsR2-integrated actinorhodin overproducing strain. The 20 gel-electrophoresis gave approximately 340 protein spots showing different protein expression patterns between these two S. lividans strains. Further MALDI-TOF analysis revealed several AfsR2-target proteins, including glyceraldehyde-3-phosphate dehydrogenase, putative phosphate transport system regulator, guanosine penta phosphate synthetase/polyribonucleotide nucleotidyltransferase, and superoxide dismutase, which suggests that the AfsR2 should be a pleiotropic regulatory protein which controls differential expressions of various kinds of genes in Streptomyces species.

Expression of Enhanced Green Fluorescent Protein from Stably Transformed Drosophila melanogaster S2 Cells

  • Lee, Jong-Min;Park, Jong-Hwa;Chung, In-Sik
    • Journal of Microbiology and Biotechnology
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    • 제10권1호
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    • pp.115-118
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    • 2000
  • Recombinant plasmids harboring a heterologous gene coding for the enhanced green fluorescent protein (EGFP) were transfected and expressed in Drosophila melanogaster S2 cells. A stable transformation of polyclonal cell populations expressing EGFP were isolated after 4 weeks of selection with hygromycin B. The recombinant EFGP expressed in transformed S2 cells consisted of a molecular weight of 27 kDa. EGFP expression was also confirmed by fluorometric measurement. The maximum EGFP concentration was about 9.3 mg/I. The present findings demonstrate not only the successful stable expression of EGFP in Drosophuila was about 9.3 mgI. The present findings demonstrate not only the successful stable expression of EGFP in Drosophila S2 cells, but also the use of EGFP as a reporter to analyze gene expression, with its potential of a Drosophila cell expression system for recombinant protein production being an alternative to a baculovirus-insect cell expression system.

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Sustained Protein Delivery System using Core/shell Nanoparticles

  • Oh, Keun-Sang;Koo, Hyoung-Mo;Yuk, Soon-Hong
    • 한국고분자학회:학술대회논문집
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    • 한국고분자학회 2006년도 IUPAC International Symposium on Advanced Polymers for Emerging Technologies
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    • pp.180-180
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    • 2006
  • A novel preparation method for core/shell nanoparticles with protein drug-loaded lipid core was designed and characterized. The lipid core is composed of lecithin and protein drug and the polymeric shell is composed of Pluronics (poly (ethylene oxide)-poly (propylene oxide)-poly(ethylene oxide) triblock copolymer, F-127 For the application of core/shell nanoparticles as a protein drug carrier, lysozyme and Vascular Endothelial Growth Factor (VEGF) were loaded into the core/shell nanoparticles by electrostatic interaction and the drug release pattern was observed by manipulating the polymeric shell.

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Preparation and Mechanical Properties of Wheat Protein Isolate Films Cross-linked with Resorcinol

  • Chandrasekhar, M.;Prabhakar, M.N.;Song, Jung-Il
    • Composites Research
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    • 제28권2호
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    • pp.40-45
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    • 2015
  • The purpose of the present work was to preparation and study of full biodegradable Eco-friendly bio-composites by using renewable resources. In this study, wheat protein isolate (WPI) films were formed by cross linking with resorcinol through solution casting method for packaging applications. By varying the resorcinol content (10, 20, 30, 40, and 50 wt %), its effect on mechanical properties of the wheat protein isolate film was measured. The addition of 20% resorcinol led to an overall increase in the tensile strength from 5.2 to 18.6 MPa and modulus increase from 780 to 1132 MPa than WPI films. The % elongation was increased from 2.8 to 9.05 when compared to unmodified WPI film. A thermal phase transition of the prepared WPI was assessed by means of DSC. FTIR is evident that the characteristic WPI spectral IR bands shifted on cross-linking with resorcinol.

포말 농축물에 의한 포말 생성의 영향인자 (The Effective Factors of n Foam Generation Using Foam Condensate)

  • 서근학;신정식;이주화
    • 한국수산과학회지
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    • 제36권5호
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    • pp.509-514
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    • 2003
  • We performed the experiment to determine the effective factors, such as the initial concentration of protein, pore size of air distributor, SAV (superficial air velocity), pH, salts and temperature related to foaming characteristics. The foam height in a foam generator was increased with the increase of the initial protein concentration and the decrease of pore size. As SAV was increased, the foam height was increased, and the optimum SAV was 0.84 cm/sec. The foam height was highest in the acid region and it was increased with the increase of salt concentration of NaCl and $NaHCO_3.$ The removal efficiencies of TSS (total suspended solid) and turbidity decreased with the increase of the initial protein concentration in the batch foam separator.

Proteomics-Driven Identification of SCO4677-Dependent Proteins in Streptomyces lividans and Streptomyces coelicolor

  • Choi, Si-Sun;Kim, Seon-Hye;Kim, Eung-Soo
    • Journal of Microbiology and Biotechnology
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    • 제20권3호
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    • pp.480-484
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    • 2010
  • AfsR2 is a global regulatory protein that stimulates antibiotic biosynthesis in both Streptomyces lividans and S. coelicolor. Previously, various afsR2-dependent genes including a putative abaA-like regulatory gene, SCO4677, were identified through comparative DNA microarray analysis. To further identify the putative SCO4677-dependent proteins, the comparative proteomics-driven approach was applied to the SCO4677-overexpressing strains of S. lividans and S. coelicolor along with the wild-type strains. The 2D gel electrophoresis gave approximately 277 protein spots for S. lividans and 207 protein spots for S. coelicolor, showing different protein expression patterns between the SCO4677-overexpressing strains and the wild-type strains. Further MALDI-TOF analysis revealed that only 18 proteins exhibited similar expression patterns in both S. lividans and S. coelicolor, suggesting that the SCO4677 could encode an abaA-like regulator that controls a few cross-species common proteins as well as many species-specific proteins in Streptomyces species.

A novel method for predicting protein subcellular localization based on pseudo amino acid composition

  • Ma, Junwei;Gu, Hong
    • BMB Reports
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    • 제43권10호
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    • pp.670-676
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    • 2010
  • In this paper, a novel approach, ELM-PCA, is introduced for the first time to predict protein subcellular localization. Firstly, Protein Samples are represented by the pseudo amino acid composition (PseAAC). Secondly, the principal component analysis (PCA) is employed to extract essential features. Finally, the Elman Recurrent Neural Network (RNN) is used as a classifier to identify the protein sequences. The results demonstrate that the proposed approach is effective and practical.

Protein Array Fabricated by Microcontact Printing for Miniaturized Immunoassay

  • Lee Woo-Chang;Lim Sang-Soo;Choi Bum-Kyoo;Choi Jeong-Woo
    • Journal of Microbiology and Biotechnology
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    • 제16권8호
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    • pp.1216-1221
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    • 2006
  • A protein array was fabricated for a miniaturized immunoassay using microcontact printing ($\mu$CP). A polydimethylsiloxane (PDMS) stamp with a 5 $\mu$m$\times$5 /$\mu$m dimension was molded from a silicon master developed by photolithography. Under optimal fabrication conditions, including the baking, incubation, and exposure time, a silicon master was successfully fabricated with a definite aspect ratio. An antibody fragment was utilized as the ink for the $\mu$CP, and transferred to an Au substrate because of the Au-thiol (-SH) interaction. The immobilization and antibody-antigen interaction were investigated with fluorescence microscopy. When human serum albumin (HSA) was applied to the protein array fabricated with an antibody against HSA, the detection limit was 100 pg/ml of HSA when using a secondary antibody labeled with a fluorescence tag. The fabricated protein array maintained its activity for 14 days.

재조합 대장균을 이용한 GFP-AFP Chimera 단백질 과량발현 및 특성 파악연구 (Overexpression of GFP-AFP Chimera Protein using Recombinant Escherichia coli and Analysis of Anti-freezing Characteristics)

  • 고지선;홍순호
    • KSBB Journal
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    • 제28권5호
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    • pp.310-314
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    • 2013
  • Antifreeze peptide from Myoxocephalus octodecemspinosus was overexpressed and purified in Escherichia coli. Green fluorescence protein-AFP chimera was constructed by integrating gfp and afp genes. Produced GFP-AFP chimera protein was purified using polyhistidine tag which was inserted at C-terminus. By addition of GFP-AFP chimera protein, freezing point of elution buffer was decreased from $-13^{\circ}C$ to $-20^{\circ}C$. This result suggested that GFP-AFP chimera can be considered as a potential candidate of novel inhibitor for gas hydrates.

Investigation of the Relationship between Protein, Message and Inducer Concentrations in Recombinant E. coli Cells

  • Jorgensen, Lene;Connor J. Thomas;Brian K. Oneill;Anton P.J. Middelbeg
    • Journal of Microbiology and Biotechnology
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    • 제7권1호
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    • pp.21-24
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    • 1997
  • Chloramphenicol acetyl transferase (CAT) protein and mRNA levels in E. coli were determined following induction of a tac::cat construct by isopropyl-${\beta}$-thiogalactopyranoside (IPTG). High cat mRNA levels did not directly reflect CAT protein levels, in either shakeflask experiments or fermentations. Furthermore, concentrations of IPTG resulting in the highest levels of expression of cat mRNA, were different to those resulting in highest levels of CAT protein. The data suggest that high transcriptional activities lead to limitations at the translational level.

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