Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Pivot Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Target Concepts:
Gene/Protein
Disease
Symptom
Drug
Enzyme
Compound
Query: EC:1.13.12.5 (
aequorin
)
1,451
document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)
Recombinant techniques are used to fuse biologically important molecules or peptides to the N-terminus of the photoprotein
aequorin
such that the binding characteristics of the molecule and the bioluminescent activity of
aequorin
are retained. This work demonstrates that the peptide region of a bulky protein can be used to develop an assay for the protein. A heterogeneous competitive binding assay was first developed for HPC4 epitope, the binding region of
protein C
, using HPC4-apoaequorin conjugate. It was observed that the binding of HPC4 epitope to its monoclonal antibody and the bioluminescence properties of
aequorin
were retained in the fusion protein. The same strategy and the same fusion protein were used to develop the assay for
protein C
. This project could potentially be a model for large biomolecules utilizing only the binding region of the protein in the labeled analyte. Also, this assay can be used in clinical diagnostics for the quantitative detection of
protein C
.
...
PMID:Using epitope-aequorin conjugate recognition in immunoassays for complex proteins. 1144 8
The protein truncation test (PTT) is important in screening for unknown mutations that cause premature termination of mRNA translation. PTT involves amplification of the interrogated sequence, in vitro transcription/translation, separation of the generated polypeptides, and detection. In this article, we report a bioluminescent protein truncation test, in which the detection of the nascent protein is performed directly in the expression mixture, within seconds, without the need for separation and purification. A DNA fragment encoding apoaequorin is fused, in-frame, downstream of the interrogated sequence. The fusion product is subjected to in vitro, coupled transcription and translation in the presence of coelenterazine. A wild-type DNA template allows translation to continue after the 3' end of the interrogated sequence, producing a chimeric protein whose C-terminal domain is the photoprotein
aequorin
. Aequorin is detected, with a high sensitivity, by its characteristic Ca(2+)-triggered, flash-type bioluminescent reaction. Active photoprotein is not produced when a truncating mutation is present in the interrogated sequence. As a model, the method was applied to the detection of truncating mutations in the
APC
gene (adenomatous polyposis coli).
...
PMID:Screening for unknown mutations by a bioluminescent protein truncation test with homogeneous detection. 2023 60
