Purify & Characterize Human Pancreatic Polypeptide Expressed in E coli

Griko, Yuri

Abstract

The region of cDNA encoding human pancreatic polypeptide (hPP) was obtained by polymerase chain reaction (PCR) and subcloned into an expression vector. The pancreatic polypeptide gene was expressed in Escherichia coli in two versions: as a cleavable fusion protein with IgG-binding synthetic ZZ domains of protein A from Staphylococcus aureus or with the 1-48 fragment of l Cro repressor. Site-specific hydrolysis by hydroxylamine was used to cleave the fusion protein, releasing the human polypeptide. The structure of the obtained hPP has been studied by scanning microcalorimetry and circular dichroism spectrometry. It has been shown that hPP in solutions close to neutral has a compact and unique spatial structure with an extended hydrophobic core. This structure is stable at 20 oC and co-operatively breaks down upon heating from this temperature.

Funding Agency

• Agency: National Institute of Health (NIH)
• Institute: National Center for Research Resources (NCRR)
• Type: Biotechnology Resource Grants (P41)
• Project #: 5P41RR004328-09
• Application #: 5224874
• Support Year: 9
• Fiscal Year: 1996

Publications

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