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Kinetic characterization of calmodulin‐troponin C chimeras binding to endothelial and neuronal nitric oxide synthases
Author(s) -
Brown Christopher C.,
Helms Katy A.,
Hill Stephanie A.,
Ghosh Dipak,
Salerno John C.,
McMurry Jonathan L.
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.984.1
Subject(s) - calmodulin , enos , troponin c , nitric oxide , endothelial nos , chemistry , angiogenesis , biochemistry , nitric oxide synthase , microbiology and biotechnology , biophysics , enzyme , biology , troponin i , medicine , organic chemistry , myocardial infarction , cancer research
Endothelial and neuronal nitric oxide synthases (eNOS and nNOS) are signal‐generating enzymes with numerous physiological effects including angiogenesis and signaling at synapses. Calmodulin (CaM) is an important regulator of NOS activity. Following on our recent report (McMurry, et al. FEBS J. 2011) describing the kinetics of CaM‐NOS binding, present results describe binding of nNOS and eNOS to Calmodulin‐Troponin C (CaM, TnC) chimeras differing in EF hand domain composition. CaM and TnC have similar structures consisting of two globular domains linked by a central helix with each domain consisting of two EF hands. Chimeras examined included each of the four domains of TnC exchanged for the analogous CaM domain. Chimeras exhibited a wide range of rate and affinity constants as determined by optical biosensing, generally reflecting their ability to activate eNOS and nNOS. Affinities ranged from subnanomolar to micromolar. Binding was rapidly dissociable in the presence of EDTA, confirming that observed binding was calcium‐dependent. This work was supported by NIH grants GM080701 and GM083317 and NSF grants 0950920 and 0922699 and a KSU Mentor‐Protégé Grant.