Linked Life Data

19121395

Source:http://linkedlifedata.com/resource/pubmed/id/19121395

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2009-2-2
pubmed:abstractText
The gene encoding acetylcholinesterase (AChE) was cloned from common carp muscle tissue. The full-length cDNA was 2368 bp that contains a coding region of 1902 bp, corresponding to a protein of 634 amino acids. The deduced amino acid sequence showed a significant homology with those of ichthyic AChEs and several common features among them, including T peptide encoded by exon T in the C-terminus. Three yeast expression vectors were constructed and introduced into the yeast Pichia pastoris. The transformant harboring carp AChE gene lacking exon T most effectively produced AChE activity extracellularly. The replacement of the native signal sequence with the yeast alpha-factor prepro signal sequence rather decreased the production. A decrease in cultivation temperature from 30 to 15 degrees C increased the activity production 32.8-fold. The purified recombinant AChE lacking T peptide, eluted as a single peak with a molecular mass of about 230 kDa on the gel filtration chromatography, exhibited the specific activity of 4970 U/mg. On the SDS-PAGE, three proteins with molecular masses of 73, 54, and 22 kDa were observed. These proteins were N-glycosylated, and their N-terminal sequence showed that the latter two were produced from the former probably by proteolytic cleavage at the C-terminal region. Thus, the recombinant AChE is homotrimer of three identical subunits with 73 kDa. The optimal temperature and pH of the recombinant were comparable to those of the native enzyme purified previously, but the values of kinetic parameters and the sensitivities to substrate inhibition and inhibitors were considerably different between them.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
1096-0279
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
64
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
205-12
pubmed:meshHeading
pubmed:year
2009
pubmed:articleTitle
Cloning and expression of carp acetylcholinesterase gene in Pichia pastoris and characterization of the recombinant enzyme.
pubmed:affiliation
Laboratory of Environmental Biochemistry, Department of Environmental Systems Engineering, Nagaoka University of Technology, Nagaoka, Niigata 940-2188, Japan.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't