Search results for "Polyhistidine-tag"
showing 3 items of 3 documents
Impact of histidine spacing on modified polyhistidine tag – Metal ion interactions
2018
Abstract Histidine rich sequences are chosen both by nature and by molecular biologists due to their high affinity towards metal ions. In this work, we examine the affinity and binding modes of Cu 2+ , Ni 2+ and Zn 2+ towards two histidine tags, the common His 6 -tag (Ac-HHHHHH-NH 2 ) and its modified sequence, which also contains six histidines, but separated with two alanine residues (Ac-HAAHAAHAAHAAHAAHAA-NH 2 ). The spatial separation of histidines has an important impact on its coordination properties. Cu 2+ and Ni 2+ complexes with Ac-HHHHHH-NH 2 are more stable than those with Ac-HAAHAAHAAHAAHAAHAA-NH 2 ; the contrary is observed for Zn 2+ . In a narrow range of pH, Cu 2+ -Ac-HHHHHH-…
Purification and analysis of polyhistidine-tagged human parvovirus B19 VP1 and VP2 expressed in insect cells
2008
Human parvovirus B19 is an autonomously replicating human pathogen with a specific tropism for human erythroid progenitor cells. There is an interest in producing empty nucleocapsids of B19 as they can be used as tools in molecular biology and diagnostics. Native B19 virus particles are formed from two structural viral proteins, VP1 and VP2. The VP2 protein alone is able to self assemble and consequently form virus-like particles (VLPs) in heterologous expression systems. Purification of recombinant VLPs has been conducted using various traditional methods. These include laborious and time-consuming, e.g. cesium chloride or sucrose gradient ultracentrifugation steps, allowing limited workin…
Expression and purification of polyhistidine-tagged firefly luciferase in insect cells
2001
The coleopteran firefly, Photinus pyralis, luciferase was produced in lepidopteran Trichoplusia ni insect cells using a baculovirus expression vector. The recombinant protein was equipped with a polyhistidine affinity tag at the carboxyl terminus and purified by immobilized metal-ion affinity chromatography in combination with an expanded bed adsorption system. This approach enabled an efficient, one-step purification protocol of a genetically modified luciferase with properties similar to those of the authentic counterpart. According to light emission measurements, the final yield of highly purified protein was 23 mg l−1 of cell culture. In addition, no specific interaction of interfering …