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- Title
Bioactivity studies of Huh - 7 cells derived human epidermal growth factor expressed in Pichia pastoris.
- Authors
Khan, Mohsin Ahmad; Ahmed, Nadeem; Khan, Muhammad Islam; Zafar, Ahmad Usman; Tahir, Saad; Anjum, Muhammad Sohail; Ali, Muhammad; Khan, Faidad; Husnain, Tayyab
- Abstract
Previously, we have reported cloning of human epidermal growth factor gene fromHuh-7cells and its extracellular expression inPichia pastoris. The presented work is a detailed report regarding molecular characterization ofHuh-7cells-derived hEGF expressed inPichia pastoriswith special reference to its glycosylation profiling and bioactivity studies. Densitometric scanning of SDS-PAGE separated extracellular proteins from hEGF recombinantPichia pastorisstrain indicated that about 84% of the extracellular proteins were glycosylated. Size exclusion chromatography using Superdex 75 prep grade column was successfully utilized to separate fractions containing glycosylated and non-glycosylated extracellular proteins. In dot blot assay, hEGF was detected in both glycosylated and non-glycosylated fractions. Bioactivity assays revealed that both glycosylated and non-glycosylated fractions were bioactive as determined by cell viability assay. It was also observed that hEGF present in non-glycosylated fraction was relatively more bioactive than hEGF present in glycosylated fraction. Bioactivity studies of Huh-7 cells-derived human epidermal growth factor expressed inPichia pastoris.
- Subjects
PICHIA pastoris; EPIDERMAL growth factor; GENE expression
- Publication
Bioscience, Biotechnology & Biochemistry, 2017, Vol 81, Issue 6, p1114
- ISSN
0916-8451
- Publication type
Article
- DOI
10.1080/09168451.2017.1295802