Results found: 3

Number of results on page

Search results

Sort By:

Limit search:

Sequence-specific Ni(II)-dependent peptide bond hydrolysis in a peptide containing threonine and histidine residues

100%

Krężel A., Mylonas M., Kopera E., Bal W.

2006

vol. 53

issue 4

721-727

Previously we demonstrated that Ni(II) complexes of Ac-Thr-Glu-Ser-His-His-Lys-NH2 hexapeptide, representing residues 120-125 of human histone H2A, and some of its analogs undergo E-S peptide bond hydrolysis. In this work we demonstrate a similar coordination and reactivity pattern in Ni(II) complexes of Ac-Thr-Glu-Thr-His-His-Lys-NH2, its threonine analogue, studied using potentiometry, electronic absorption spectroscopy and HPLC. For the first time we present the detailed temperature and pH dependence of such Ni(II)-dependent hydrolysis reactions. The temperature dependence of the rate of hydrolysis yielded activation energy Ea = 92.0 kJ mol-1 and activation entropy ΔS≠ = 208 J mol-1 K-1. The pH profile of the reaction rate coincided with the formation of the four-nitrogen square-planar Ni(II) complex of Ac-Thr-Glu-Thr-His-His-Lys-NH2. These results expand the range of protein sequences susceptible to Ni(II) dependent cleavage by those containing threonine residues and permit predictions of the course of this reaction at various temperatures and pH values.

Overexpression of genes involved in phytochelatin biosynthesis in Escherichia coli: effects on growth, cadmium accumulation and thiol level.

76%

Wawrzyńska A., Wawrzyński A., Gaganidze D., Kopera E., Piątek K., Bal W., Sirko A.

2005

vol. 52

issue 1

109-116

In Escherichia coli, heterologous production of Schizosaccharomyces pombe phytochelatin synthase (PCS) along with overproduction of E. coli serine acetyltransferase (SAT) and γ-glutamylcysteine synthase (γECS) was achieved and resulted in the accumulation of phytochelatins in bacterial cells. Overproduction of either γECS alone or simultaneous production of all three proteins in bacterial cells were accompanied by reduced growth rate in liquid cultures. Interestingly, bacteria overproducing either γECS or both SAT and γECS (with elevated level of γ-glutamylcysteine but not of phytochelatins) were able to accumulate more cadmium per dry weight than the control. However, the most efficient cadmium accumulation was observed in bacteria with elevated levels of all three proteins: SAT, γECS and PCS. Therefore, "pushing" the entire pathway might be the most promising approach in modification of bacteria for potential bioremediation purposes because the level of intermediates, cysteine and glutathione, can limit the rate of production of phytochelatins. However, in such bacteria other metabolic process might become limiting for efficient growth.

Expression, purification and characterization of glycosylated influenza H5N1 hemagglutinin produced in Pichia pastoris

64%

Kopera E., Dwornyk A., Kosson P., Florys K., Sączyńska V., Dębski J., Cecuda-Adamczewska V., Szewczyk B., Zagórski-Ostoja W., Grzelak K.

2014

vol. 61

issue 3

597-602

The A/swan/Poland/305-135V08/2006 (H5N1-subtype) hemagglutinin (HA) gene was cloned and expressed in yeast Pichia pastoris (P. pastoris). The HA cDNA lacking the C-terminal transmembrane anchor-coding sequence was fused to an α-factor leader peptide and placed under control of the methanol-inducible P. pastoris alcohol oxidase 1 (AOX1) promoter. Two P. pastoris strains: SMD 1168 and KM 71 were used for protein expression. Recombinant HA protein was secreted into the culture medium reaching an approximately 15 mg/L (KM 71 strain). Fusion protein with a His6 tag was purified to homogeneity in one step affinity chromatography. SDS-PAGE and MS/MS analysis indicated that the protein is cleaved into HA1 and HA2 domains linked by a disulfide bond. Analysis of the N-linked glycans revealed that the overexpressed HA is fully glycosylated at the same sites as the native HA in the vaccine strain. Immunological activity of the hemagglutinin protein was tested in mice, where rHA elicited a high immune response.

Refine search results

3 Acta Biochimica Polonica

1 2014

1 2006

1 2005

3 Kopera E.

2 Bal W.

1 Cecuda-Adamczewska V.

1 Dwornyk A.

1 Dębski J.

1 Florys K.

1 Gaganidze D.

1 Grzelak K.

1 Kosson P.

1 Krężel A.

1 Mylonas M.

1 Piątek K.

1 Sirko A.

1 Szewczyk B.

1 Sączyńska V.

1 Wawrzyńska A.

1 Wawrzyński A.

1 Zagórski-Ostoja W.

Search results

Sequence-specific Ni(II)-dependent peptide bond hydrolysis in a peptide containing threonine and histidine residues

Overexpression of genes involved in phytochelatin biosynthesis in Escherichia coli: effects on growth, cadmium accumulation and thiol level.

Expression, purification and characterization of glycosylated influenza H5N1 hemagglutinin produced in Pichia pastoris