Linked Life Data

7827501

Source:http://linkedlifedata.com/resource/pubmed/id/7827501

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
1995-2-21
pubmed:abstractText
Recombinant Herpes Simplex Virus Type 1 thymidine kinase (TK) was isolated in a fast and gentle two-step procedure from Escherichia coli as a thrombin cleavable fusion protein. The TK was expressed as an inducible glutathione S-acetyl transferase fusion protein and purified in a first step by glutathione affinity chromatography. Proteolytic cleavage of the column bound TK with thrombin led to a truncated enzyme, resulting from two new and hitherto unknown cleavage sites, determined by N-terminal sequencing. In a second step, the TK was further purified from the cleavage products by ATP affinity chromatography, yielding homogeneously pure TK as shown by SDS-PAGE and mass spectrometry. Both the fusion protein and the purified enzyme show enzymatic activity with the same Km value of 0.2 microM for the natural substrate thymidine. Determination of the native molecular weight indicated that the pure enzyme and the fusion protein are biologically active as homodimers. Therefore the recombinant enzyme has the same biochemical characteristics as the viral TK, expressed in infected cells.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
1046-5928
pubmed:author
pubmed:issnType
Print
pubmed:volume
5
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
432-41
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
1994
pubmed:articleTitle
A fast method for obtaining highly pure recombinant herpes simplex virus type 1 thymidine kinase.
pubmed:affiliation
Department of Pharmacy, Swiss Federal Institute of Technology, ETH-Zentrum, Zürich.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't