Linked Life Data

8340354

Source:http://linkedlifedata.com/resource/pubmed/id/8340354

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
1993-9-2
pubmed:abstractText
A glycosylphosphatidylinositol (GPI)-anchored protein, 5'-nucleotidase [EC 3.1.3.5], was released from the membrane of bovine liver by use of phosphatidylinositol-specific phospholipase C (PI-PLC) of Bacillus thuringiensis and purified by several column chromatographies to a homogeneous state. The purified protein has an apparent molecular mass of 61 kDa, as estimated by SDS-polyacrylamide gel electrophoresis. From the partial amino acid sequence of a tryptic peptide, mixed oligonucleotides were synthesized and used to screen a lambda gt11 liver cDNA library, and one positive clone, pE1, was isolated. Since the insert of the clone lacked the NH2-terminal coding region, another lambda gt11 liver cDNA library was screened by using a synthetic probe corresponding to the 5' region of the insert of pE1. Three additional cDNA clones were obtained. Sequencing of these cDNAs revealed an open reading frame that encodes a 574-residue polypeptide with a calculated mass of 63,084 Da. The predicted structure showed two highly hydrophobic stretches at both ends of the protein, like those of rat and human 5'-nucleotidases. The NH2-terminal 26 residues comprise a signal peptide and the COOH-terminal hydrophobic stretch may serve as a signal for the posttranslational GPI modification. An expression vector of the cDNA, pSVNT, was constructed in a mammalian expression vector pSVL and the 5'-nucleotidase activity was transiently expressed in COS-1 cells. The expressed activity was about 8 times higher than the pSVL-transfected control activity. PI-PLC released 45% of the transiently expressed 5'-nucleotidase activity, indicating that the cDNA isolated here encodes this enzyme expressed as a GPI-anchored protein.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0021-924X
pubmed:author
pubmed:issnType
Print
pubmed:volume
113
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
607-13
pubmed:dateRevised
2007-12-19
pubmed:meshHeading
pubmed:year
1993
pubmed:articleTitle
Purification and cDNA cloning of bovine liver 5'-nucleotidase, a GPI-anchored protein, and its expression in COS cells.
pubmed:affiliation
Department of Microbial Chemistry, Faculty of Pharmaceutical Sciences, Nagoya City University.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't