@article{
 title = {An alternative assay to discover potential calmodulin inhibitors using a human fluorophore-labeled CaM protein},
 type = {article},
 year = {2009},
 identifiers = {[object Object]},
 volume = {387},
 id = {28a06b39-eb92-3661-ad83-00e4888c8f2c},
 created = {2017-08-31T00:54:41.545Z},
 file_attached = {false},
 profile_id = {7da96f2b-cf05-3448-83f2-2c1e91a65514},
 last_modified = {2017-08-31T00:57:51.591Z},
 read = {false},
 starred = {false},
 authored = {true},
 confirmed = {false},
 hidden = {false},
 private_publication = {false},
 abstract = {This article describes the development of a new fluorescent-engineered human calmodulin, hCaM M124C-mBBr, useful in the identification of potential calmodulin (CaM) inhibitors. An hCaM mutant containing a unique cysteine residue at position 124 on the protein was expressed, purified, and chemically modified with the fluorophore monobromobimane (mBBr). The fluorophore-labeled protein exhibited stability and functionality to the activation of calmodulin-sensitive cAMP phosphodiesterase (PDE1) similar to wild-type hCaM. The hCaM M124C-mBBr is highly sensitive to detecting inhibitor interaction given that it showed a quantum efficiency of 0.494, approximately 20 times more than the value for wild-type hCaM, and a large spectral change (∼80% quenching) when the protein is in the presence of saturating inhibitor concentrations. Two natural products previously shown to act as CaM inhibitors, malbrancheamide (1) and tajixanthone hydrate (2), and the well-known CaM inhibitor chlorpromazine (CPZ) were found to quench the hCaM M124C-mBBr fluorescence, and the IC 50  values were comparable to those obtained for the wild-type protein. These results support the use of hCaM M124C-mBBr as a fluorescence biosensor and a powerful analytical tool in the high-throughput screening demanded by the pharmaceutical and biotechnology industries. © 2009 Elsevier Inc. All rights reserved.},
 bibtype = {article},
 author = {González-Andrade, M. and Figueroa, M. and Rodríguez-Sotres, R. and Mata, R. and Sosa-Peinado, A.},
 journal = {Analytical Biochemistry},
 number = {1}
}

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An hCaM mutant containing a unique cysteine residue at position 124 on the protein was expressed, purified, and chemically modified with the fluorophore monobromobimane (mBBr). The fluorophore-labeled protein exhibited stability and functionality to the activation of calmodulin-sensitive cAMP phosphodiesterase (PDE1) similar to wild-type hCaM. The hCaM M124C-mBBr is highly sensitive to detecting inhibitor interaction given that it showed a quantum efficiency of 0.494, approximately 20 times more than the value for wild-type hCaM, and a large spectral change (∼80% quenching) when the protein is in the presence of saturating inhibitor concentrations. Two natural products previously shown to act as CaM inhibitors, malbrancheamide (1) and tajixanthone hydrate (2), and the well-known CaM inhibitor chlorpromazine (CPZ) were found to quench the hCaM M124C-mBBr fluorescence, and the IC 50 values were comparable to those obtained for the wild-type protein. 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