An Efficient System for the Expression and Purification of Yeast Geranylgeranyl Protein Transferase Type I

To purify the geranylgeranyl protein transferase type I (GGPT-I) efficiently, a gene expression system using the pGEX-4T-1 vector was constructed. The cal 1 gene, encoding the β subunit of GGPT-I, was subcloned into the pGEX-4T-1 vector and co-transformed into E. coli cells harboring the ram2 gene,...

Full description

Saved in:
Bibliographic Details
Published inBMB reports Vol. 31; no. 1; pp. 77 - 82
Main Authors HyunKyung Kim, Youngah Kim, Chul Hak Yang
Format Journal Article
LanguageKorean
Published 생화학분자생물학회 30.01.1998
Online AccessGet full text

Cover

More Information
Summary:To purify the geranylgeranyl protein transferase type I (GGPT-I) efficiently, a gene expression system using the pGEX-4T-1 vector was constructed. The cal 1 gene, encoding the β subunit of GGPT-I, was subcloned into the pGEX-4T-1 vector and co-transformed into E. coli cells harboring the ram2 gene, the α subunit gene of GGPT-I. GGPT-I was highly expressed as a fusion protein with glutathione S-transferase (GST) in E. coli, purified to homogeneity by glutathione-agarose affinity chromatography, and the GST moiety was excised by thrombin treatment. The purified yeast GGPT-I showed a dose-dependent increase in the transferase activity, and its apparent K_m value for an undecapeptide fused with GST (GST-PEP) was 0.66μM and the apparent K_m value for geranylgeranyl pyrophosphate (GGPP) was 0.071 μM.
Bibliography:Korean Society for Biochemistry and Molecular Biology
ISSN:1976-6696