Journal of Microbiology

Expression of human hypoxanthine phosphoribosyltransferase in Insect Cells Using a Baculovirus Vector: Lee, Chong Ho , Yang, Ji Won , Baek, Sang Ki , Yang, jai Myung; J. Microbiol. 1995;33(1):85-90.

Abstract: The hypoxanthine phosphoribosyltransferase (HPRT) catalyzes the conversion of hypoxanthine and guanine to the mononucleotide, IMP and GMP, respectively. For construction of recombinant AcNPV carrying human HPRT, a transfer vector p918 constructed by cloning full-length cDNA for human HPRT into pVL1393 and AnNPV genomic DNA were co-transfected into Sf21 cells. The tissue culture fluid containing extracellular virus was plaque assayed and a recombinant virus with occlusion minus phenotype was obtained by three rounds of plaque purification. Southern blot analysis and PRC results confirmed the insertion of the human HPRT cDNA within the recombinant virus(AcHPRT918). SDS-PAGE and Western blot analysis of the Sf21 cell extracts infected with AcHPRT918 indicated that human HPRT was expressed in insect cell. Large quantities of functional HPRT expressed in insect cells would facilitate characterization of the biological properties of this enzyme.

Regulation of Glycogen Concentration by the Histidine-Containing Phosphocarrier Protein HPr in Escherichia coli: Byoung-Mo Koo , Yeong-Jae Seok; J. Microbiol. 2001;39(1):24-30.

Abstract: In addition to effecting the catalysis of sugar uptake, the bacterial phosphoenolpyruvate:sugar phosphotransferase system regulates a variety of physiological processes. In a previous paper [Seok et al., (1997) J. Biol. Chem. 272, 26511-26521], we reported the interaction with and allosteric regulation of Escherichia coli glycogen phosphorylase activity by the histidine-containing phosphocarrier protein HPr in vitro. Here, we show that the specific interaction between HPr and glycogen phosphorylase occurs in vivo. To address the physiological role of the HPr-glycogen phosphorylase complex, intracellular glycogen levels were measured in E. coli strains transformed with various plasmids. While glycogen accumulated during the transition between exponential and stationary growth phases in wildtype cells, it did not accumulate in cells overproducing HPr or its inactive mutant regardless of the growth stage. From these results, we conclude that HPr mediates crosstalk between sugar uptake through the phosphoenolpyruvate:sugar phosphotransferase system and glycogen breakdown. The evolutionary significance of the HPr-glycogen phosphorylase complex is suggested.

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