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Modification of human neuron‐specific enolase for application to radioimmunoassay
Author(s) -
Aoki Takashi,
Kaneta Mitsuhiro,
Onagi Hitoshi,
Morikawa Junji,
Tsubota Nobuyuki,
Watabe Hiroyuki
Publication year - 1996
Publication title -
iubmb life
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.132
H-Index - 113
eISSN - 1521-6551
pISSN - 1521-6543
DOI - 10.1080/15216549600201511
Subject(s) - enolase , histidine , tyrosine , biochemistry , recombinant dna , radioimmunoassay , chemistry , residue (chemistry) , specific activity , escherichia coli , chromatography , microbiology and biotechnology , biology , enzyme , immunohistochemistry , gene , immunology
A recombinant human neuron‐specific enolase (R‐NSE), isolated from Escherichia coli, could not be used in an RIA system because of instability upon labeling. To apply R‐NSE to RIA and to simplify the purification procedure, the N‐ and C‐terminals of R‐NSE were modified by tyrosine‐ and histidine‐tagging, respectively. SY‐NSE, containing one additional tyrosine residue, was obtained from both soluble and insoluble fractions. More derivatives tagged by two or four tyrosine residues were expressed, but only in the insoluble fraction. SY‐NSE and SY‐NSE.H6 (containing six histidine residues at C‐terminal of SY‐NSE) purified from the soluble fraction were applicable to the RIA system, indicating that the addition of a tyrosine residue at the terminal is effective if the antigen is unstable during labeling.