A monoclonal antibody that distinguishes phospho- and dephosphorylated forms of cardiac troponin-I

Cummins, Bernadette; Russell, Geoffrey J; Cummins, Peter L.

doi:10.1042/bst019161s

Cited by 5 publications

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“…9 The protein itself contains cysteine residues capable of producing intramolecular disulphide bridges and can also be phosphorylated, resulting in molecular forms that are distinguishable using immunobinding techniques. 10 Antibody interactions can occur between free or complexed cTnI, as well as between phosphorylated, dephosphorylated, reduced and oxidiz ed forms. In addition to these forms, enzyme degradation products and high molecular weight covalent complexes may interact with antibodies.…”

Section: Introductionmentioning

confidence: 99%

Measurement of cardiac troponin I in striated muscle using three experimental methods

et al. 2003

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Background Qualitative and quantitative measures of cardiac troponin I (cTnI) in striated muscle have been reported as part of diverse investigations. However, there is disparity in the literature regarding the findings of these analyses. The cTnI molecule can exist in phosphorylated, non-phosphorylated, reduced, non-reduced, complexed or non-complexed forms. Each of these forms can change the antigenicity of cTnI, resulting in different antibody-antigen interactions in different experimental formats, thereby giving rise to the disparities in the literature.

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Section: Introductionmentioning

confidence: 99%

Measurement of cardiac troponin I in striated muscle using three experimental methods

et al. 2003

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The use of phosphate‐affinity SDS‐PAGE to measure the cardiac troponin I phosphorylation site distribution in human heart muscle

Messer

Gallon

McKenna

et al. 2009

Proteomics Clinical Apps

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We have used phosphate affinity SDS-PAGE to separate the phosphorylated species of cardiac troponin I (cTnI). To test the method we phosphorylated pure cTnI with protein kinase A catalytic subunit and observed up to six bands corresponding to 0, 1P, 2P, 3P, 4P and 5P phospho-species. We examined the phospho-species of cTnI in human heart myofibrillar extracts by phosphate affinity SDS-PAGE and Western blotting with a non-specific troponin I (TnI) antibody. In donor heart samples the bis-phosphorylated species of cTnI predominated and no more highly phosphorylated species were not detectable (0P was 10.3±1.9%, 1P, 17.5±3.5%, 2P, 72.2±4.7%, 11 samples). Total phosphorylation was 1.62±0.06 molsPi/mol TnI. In myofibrils from end-stage failing hearts, the unphosphorylated cTnI species predominated (0P was 78.5±1.8%, 1P, 17.5±1.9%, 2P, 4.0±0.7%, total phosphorylation 0.26±0.02 molsPi/mol TnI, five samples). Muscle from patients with hypertrophic obstructive cardiomyopathy was also largely unphosphorylated (0P was 76.6±3.1%, 1P, 17.5±2.7%, 2P, 5.9±0.8%, total phosphorylation 0.29±0.04 molsPi/mol TnI, 19 samples). Using a range of phospho-specific antibodies we demonstrated that 3/4 of the bis-phosphorylated band of donor heart cTnI is phosphorylated at Ser22 and Ser23 in approximately equal amounts and that phosphorylation of Ser43 and Thr142 was not detected.

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Role of regulatory proteins (troponin-tropomyosin) in pathologic states

Malhotra

1994

Mol Cell Biochem

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In vertebrate striated muscle, troponin-tropomyosin is responsible, in part, not only for transducing the effect of calcium on contractile protein activation, but also for inhibiting actin and myosin interaction when calcium is absent. The regulatory troponin (Tn) complex displays several molecular and calcium binding variations in cardiac muscles of different species and undergoes genetic changes with development and in various pathologic states. Extensive reviews on the role of tropomyosin (Tm) and Tn in the regulation of striated muscle contraction have been published describing the molecular mechanisms involved in contractile protein regulation. In our studies, we have found an increase in Mg2+ ATPase activity in cardiac myofibrils from dystrophic hamsters and in rats with chronic coronary artery narrowing. The abnormalities in myofibrillar ATPase activity from cardiomyopathic hamsters were largely corrected by recombining the preparations with a TnTm complex isolated from normal hamsters indicating that the TnTm may play a major role in altered myocardial function. We have also observed down regulation of Ca2+ Mg2+ ATPase of myofibrils from hypertrophic guinea pig hearts, myocardial infarcted rats and diabetic-hypertensive rat hearts. In myosin from diabetic rats, this abnormality was substantially corrected by adding troponin-tropomyosin complex from control hearts. All of these disease models are associated with decreased ATPase activities of pure myosin and in the case of rat and hamster models, shifts of myosin heavy chain from alpha to beta predominate. In summary, there are three main troponin subunit components which might alter myofibrillar function however, very few direct links of molecular alterations in the regulatory proteins to physiologic and pathologic function have been demonstrated so far.

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A monoclonal antibody that distinguishes phospho- and dephosphorylated forms of cardiac troponin-I

Cited by 5 publications

References 0 publications

Measurement of cardiac troponin I in striated muscle using three experimental methods

Measurement of cardiac troponin I in striated muscle using three experimental methods

The use of phosphate‐affinity SDS‐PAGE to measure the cardiac troponin I phosphorylation site distribution in human heart muscle

Role of regulatory proteins (troponin-tropomyosin) in pathologic states

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