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Generation of engineered recombinant hepatocyte growth factor cleaved and activated by Genenase I
- Source :
- Journal of Biotechnology. 133(4):478-485
- Publication Year :
- 2008
- Publisher :
- Elsevier, 2008.
-
Abstract
- 金沢大学がん研究所附属分子標的がん医療研究開発センター<br />Hepatocyte growth factor (HGF) is biosynthesized as a biologically inactive, single-chain form (pro-HGF). Its activation is associated with cleavage at Arg494-Val495 into a two-chain mature form composed of disulfide-linked α- and β-chains. Because serum is a major source of HGF activator (the predominant serine protease responsible for the processing of pro-HGF), serum-free production of recombinant, two-chain HGF had not been established. In this study, to enable serum-free production of two-chain HGF, we generated engineered human pro-HGFs that can be specifically cleaved and activated by Genenase I. Since Genenase I specifically cleaves the C-terminus of the His-Tyr sequence, which does not exist in human HGF, Arg494 (the C-terminus of the HGF α-chain) was replaced by His-Tyr, Ala-Ala-His-Tyr, Pro-Gly-His-Tyr, or Pro-Gly-Ala-Ala-His-Tyr. Genenase I cleaved engineered pro-HGFs specifically at the replaced amino acid sequences, forming a disulfide-linked two-chain form. The cleavage was most efficient in the case of the Pro-Gly-Ala-Ala-His-Tyr sequence, and cleaved HGFs displayed biological activities identical to those of wild-type HGF. Considering a potential medical application of HGF, the present technique is valuable because it enables the production of recombinant, two-chain HGF entirely without serum and extends the choice of host cells and organisms for recombinant production. © 2007 Elsevier B.V. All rights reserved.
- Subjects :
- Serum
medicine.medical_treatment
Blotting, Western
Molecular Sequence Data
Activation
Bioengineering
Spodoptera
Cleavage (embryo)
Protein Engineering
Applied Microbiology and Biotechnology
Models, Biological
law.invention
Cell Line
law
medicine
Animals
Humans
Amino Acid Sequence
Protein Precursors
Genenase I
Recombinant Hepatocyte Growth Factor
Serine protease
chemistry.chemical_classification
Hepatocyte growth factor
biology
Growth factor
Serine Endopeptidases
Subtilisin
General Medicine
Recombinant Proteins
Amino acid
medicine.anatomical_structure
Biochemistry
chemistry
Hepatocyte
biology.protein
Recombinant DNA
Electrophoresis, Polyacrylamide Gel
Biotechnology
medicine.drug
Subjects
Details
- Language :
- English
- ISSN :
- 01681656
- Volume :
- 133
- Issue :
- 4
- Database :
- OpenAIRE
- Journal :
- Journal of Biotechnology
- Accession number :
- edsair.doi.dedup.....89cadb8566b5f33a01493749e7fd4252