Constitutive Optimized Production of Streptokinase in Saccharomyces cerevisiae Utilizing Glyceraldehyde 3-Phosphate Dehydrogenase Promoter of Pichia pastoris.
A novel expression vector constructed from genes of Pichia pastoris was applied for heterologous gene expression in Saccharomyces cerevisiae. Recombinant streptokinase (SK) was synthesized by cloning the region encoding mature SK under the control of glyceraldehyde 3-phosphate dehydrogenase (GAP) pr...
| Publicado en: | BioMed Research International Vol. 2013; pp. 268249 - 268250 |
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| Autores principales: | , , , , |
| Formato: | Journal Article |
| Publicado: |
Wiley-Blackwell
2013
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| Acceso en línea: | Ver este registro en EBSCOhost |
| fields | @attributes: recordID: 1 pdfLink: plink: https://search.ebscohost.com/login.aspx?direct=true&db=ccm&AN=104108789&site=ehost-live header: @attributes: shortDbName: ccm uiTerm: 104108789 longDbName: CINAHL Complete uiTag: AN controlInfo: bkinfo: dissinfo: jinfo: jid: 23146133 FT2T jtl: BioMed Research International issn: 23146133 maglogo: N pubinfo: dt: 2013 vid: 2013 pid: 480 pub: Wiley-Blackwell place: Malden, Massachusetts artinfo: ui: 104108789 2012354666 NLM24171161 PMC3793500 104108789 ppf: 268249 ppct: 1 formats: fmt: @attributes: type: P tig: atl: Constitutive Optimized Production of Streptokinase in Saccharomyces cerevisiae Utilizing Glyceraldehyde 3-Phosphate Dehydrogenase Promoter of Pichia pastoris. aug: au: Vellanki, Ravi N Potumarthi, Ravichandra Doddapaneni, Kiran K Anubrolu, Naveen Mangamoori, Lakshmi N affil: Centre for Biotechnology, Institute of Science and Technology, Jawaharlal Nehru Technological University, Kukatpally, Hyderabad, Andhra Pradesh 500085, India ; Ontario Cancer Institute, Princess Margaret Cancer Centre, 610 University Avenue, Toronto, ON, Canada M5G 2M9. sug: subj: Oxidoreductases Peptides Yeasts Genes Yeasts Metabolism Streptokinase Carbon Metabolism Enzymes Fibrin Metabolism Nitrogen Metabolism Fibrinolytic Agents Metabolism Genetics ab: A novel expression vector constructed from genes of Pichia pastoris was applied for heterologous gene expression in Saccharomyces cerevisiae. Recombinant streptokinase (SK) was synthesized by cloning the region encoding mature SK under the control of glyceraldehyde 3-phosphate dehydrogenase (GAP) promoter of Pichia pastoris in Saccharomyces cerevisiae. SK was intracellularly expressed constitutively, as evidenced by lyticase-nitroanilide and caseinolytic assays. The functional activity was confirmed by plasminogen activation assay and in vitro clot lysis assay. Stability and absence of toxicity to the host with the recombinant expression vector as evidenced by southern analysis and growth profile indicate the application of this expression system for large-scale production of SK. Two-stage statistical approach, Plackett-Burman (PB) design and response surface methodology (RSM) was used for SK production medium optimization. In the first stage, carbon and organic nitrogen sources were qualitatively screened by PB design and in the second stage there was quantitative optimization of four process variables, yeast extract, dextrose, pH, and temperature, by RSM. PB design resulted in dextrose and peptone as best carbon and nitrogen sources for SK production. RSM method, proved as an efficient technique for optimizing process conditions which resulted in 110% increase in SK production, 2352 IU/mL, than for unoptimized conditions. pubtype: Academic Journal doctype: Journal Article ougenre: Article language: English refInfo: holdings: @attributes: islocal: N |
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