Efficient expression and purification of human interferon alpha2b in the methylotrophic yeast, Pichia pastoris

Shi, Linmei; Wang, Dongming; Chan, Weiguang; Cheng, Luzi

doi:10.1016/j.pep.2007.03.005

Cited by 27 publications

(24 citation statements)

References 18 publications

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“…This feature makes IFN effective in the treatment of some chronic and infectious diseases. Recombinant IFN with high biological activity has been obtained at *10 8 -10 9 IU mg -1 in E. coli (Srivastava et al 2005) and Pichia pastoris (Shi et al 2007) but complicated purification procedures and high-dose injection are needed when IFN is used clinically. An orally-delivered transgenic potato system was also employed to express IFN, but the activity of this recombinant IFN was very low (only 567 IU g -1 tissue) (Ohya et al 2001).…”

Section: Biological Activity Of Ifn Produced By L Lactismentioning

confidence: 99%

“…Several host-vector systems have been used to express IFN, including Escherichia coli (Srivastava et al 2005), Pichia pastoris (Shi et al 2007) and transgenic potato plants (Ohya et al 2001). To offer an additional food-grade expression system, Lactococcus lactis, a 'generally regarded as safe' (GRAS) microorganism, was chosen as the host to produce IFN in our previous study.…”

Section: Introductionmentioning

confidence: 99%

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Improvement of human interferon alpha secretion by Lactococcus lactis

et al. 2010

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To improve the secretion and expression of human interferon alpha 2b (IFN) in Lactococcus lactis, a synthetic pro-peptide, LEISSTCDA (LEISS), was fused to the N-terminus of IFN. This gave a higher secretion efficiency (12% vs. 5%) and yield (*2.8-fold) of IFN. The signal peptide, SP SlpA (SlpA, an S-layer protein of Lactobacillus brevis), was also tested to secrete IFN instead of SP Usp45 (Usp45, the main secreted protein in L. lactis). This gave increased IFN secretion (*3-fold) but lower total production. All the recombinant IFN had appropriate bioactivities in an antiviral assay.

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Section: Biological Activity Of Ifn Produced By L Lactismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Improvement of human interferon alpha secretion by Lactococcus lactis

et al. 2010

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“…Other reports used GS115 strain which has mutation in the histidinol dehydrogenase gene (his4) that prevents it to synthesize histidine. So, It allow the selection of expression vectors containing HIS4 upon transformation (Shi et al, 2007;Ghosalkar et al, 2008;Li et al, 2007). Our rhIFNα-2b was produced as extracellular protein with 24.05 kDa in size containing polyhistidine tag and c-myc epitope in its C terminus.…”

Section: Introductionmentioning

confidence: 99%

Antiproliferative Activity of Recombinant Human Interferon Alpha2b on Estrogen Positive Human Breast Cancer McF-7 Cell Line

Ningrum

Wisnuwardhani

Santoso

et al. 2015

Indonesian J. Pharm.

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Indonesia is the top three countries with hepatitis and moreover has 40.000 cancer mortalities per year. Interferon alpha 2b (IFNα-2b) is a therapeutic standard for cancer and hepatitis B/C treatments. We developed recombinant human interferon alpha 2b (rhIFNα-2b) in methilotropic yeast Pichia pastoris X-33. The protein was produced as extracellular protein with 24 kDa in size. This research was aimed to characterize the protein based on its amino acid sequence and to study its antiproliferative activity on MCF-7 cell line. Amino acid sequencing by using MALDI TOF TOF mass spectrometry with trypsin as proteolytic enzyme identified protein as hIFNα-2b with 33% of amino acid coverage. The antiproliferative activity was determined by 3-[4.5-dimethyltiazol-2il]-2.5-diphenylltetrazolium bromide (MTT) assay. Based on several studies about the synergistic activity of rhIFNα-2b with other anticancer drugs, we combined our rhIFNα-2b with tamoxifen (tmx). The growth percentage of the cells after being treated with 1µM of tmx at various concentrations of rhIFNα-2b was compared with that of untreated cell. This study showed that the antiproliferative activity was dose-dependently. Cell viability assay with calcein and ethidium bromide-based staining by fluorescence microscope confirmed that the rhIFNα-2b had ability to inhibit proliferation of human breast cancer cell line MCF-7.

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“…It is apparent from Table 1 [124][125][126][127][128][129][130][131][132][133][134][135][136][137][138][139][140][141][142][143], that all 20 rPRTs are derived from four sources, yeast (11), insects (4) and CHO (4), and egg (1). Each purification scheme shown here, like those described in Sections 3.3-3.7 before, contains one, two, three, or even more LC steps in combination with other non-LC separation methods.…”

Section: Order Of a Series Of Liquid Chromatographymentioning

confidence: 99%