ABSTRACT
Cardiac troponin (cTn) is made up of three subunits, cTnC, cTnI, and cTnT. The regulatory N-terminal domain of cTnC (cNTnC) controls cardiac muscle contraction in a calcium-dependent manner. We show that calcium-saturated cNTnC can adopt two different orientations, with the "active" orientation consistent with the 2020 cryo-EM structure of the activated cardiac thin filament by Yamada et al. Using solution NMR 15N R2 relaxation analysis, we demonstrate that the two domains of cTnC tumble independently (average R2 10 s-1), being connected by a flexible linker. However, upon addition of cTnI1-77, the complex tumbles as a rigid unit (R2 30 s-1). cTnI phosphomimetic mutants S22D/S23D, S41D/S43D and dilated cardiomyopathy- (DCM-)associated mutations cTnI K35Q, cTnC D75Y, and cTnC G159D destabilize the active orientation of cNTnC, with intermediate 15N R2 rates (R2 17-23 s-1). The active orientation of cNTnC is stabilized by the flexible tails of cTnI, cTnI1-37 and cTnI135-209. Surprisingly, when cTnC is incorporated into complexes lacking these tails (cTnC-cTnI38-134, cTnC-cTnT223-288, or cTnC-cTnI38-134-cTnT223-288), the cNTnC domain is still immobilized, revealing a new interaction between cNTnC and the IT-arm that stabilizes a "dormant" orientation. We propose that the calcium sensitivity of the cardiac troponin complex is regulated by an equilibrium between active and dormant orientations, which can be shifted through post-translational modifications or DCM-associated mutations.
Subject(s)
Cardiomyopathy, Dilated/genetics , Mutation , Myocardium/metabolism , Troponin C/genetics , Calcium/metabolism , Cardiomyopathy, Dilated/metabolism , Humans , Magnetic Resonance Spectroscopy , Models, Molecular , Multiprotein Complexes/chemistry , Multiprotein Complexes/metabolism , Phosphorylation , Protein Binding , Protein Domains , Tropomyosin/chemistry , Tropomyosin/metabolism , Troponin C/chemistry , Troponin C/metabolism , Troponin I/chemistry , Troponin I/metabolism , Troponin T/chemistry , Troponin T/metabolismABSTRACT
Clobazam (CLB) is a commonly used oral antiepileptic drug (AED) that has been shown to be effective in various forms of epilepsy. Given its distinct 1,5-benzodiazepine structure, rapid absorption, minimal drug interactions, and favorable safety profile, CLB displays unique properties when compared to other commonly used benzodiazepines. Recent evidence has shown that CLB may demonstrate therapeutic efficacy in status epilepticus (SE). The objective of this systematic review was to summarize the available evidence pertaining to the efficacy of CLB use in SE. An electronic literature search of Medline (1946 to November 6, 2017), Embase (1974 to November 6, 2017), and the Cochrane Library (1999 to November 6, 2017) databases was performed to identify reports of CLB use in SE. After screening and full text review, a total of 15 articles were included: 8 retrospective studies, 2 case series, and 5 case reports. Efficacy rates for CLB have varied among reports. Overall, based on the retrospective studies, a total of 76 patients with SE have been reported. CLB was introduced within 2-4 days from SE onset and has been reported to contribute to remission in 36 patients (47%). CLB maintenance dose ranged from 10 to 60 mg/day. However, the results need to be interpreted carefully because SE patients are a heterogeneous group with different etiologies and disease severities, and the response to CLB might vary in different patient population or seizure types. In conclusion, there is not sufficient evidence to determine the safety and efficacy of clobazam in the setting of SE. However, the current limited evidence combined with the unique characteristics of CLB suggest that the drug might be considered as an add-on option in SE patients, with a suggested dosage range of 10-60 mg/day. Prospective studies are needed to fully establish the role of CLB in the management of SE.
