Troponin replacement in permeabilized cardiac muscle Reversible extraction of troponin I by incubation with vanadate

FEBS Letters

Strauss, John D.¹  Troschka, Monika¹  Bletz, Christel¹  Zeugner, Claudia¹  Rüegg, J.Caspar¹  Van Eyk, Jennifer E.¹ 

[1] II Physiologisches Institut, Universität Heidelberg, INF 326, 6900 Heidelberg 1, Germany

关键词: Troponin;    Troponin I;    Vanadate;    Contractility;    Muscle regulation;    TnI;    troponin;    TnC;    troponin C;    Tnt;    troponin T;    DOC-TCA;    deoxycholate trichloroacetic acid;    ATP;    adenosine triphosphate;    EDTA;    ethylene-diamine tetraacetic acid;    EGTA;    ethylene glycol (bis(β-aminoethyl ether) N;    N;    N″;    N″ tetraacetic acid;    DTE;    dithioerythritol;    NADH;    nicotinamide adenine dinucleotide;    UV;    ultraviolet;    HRP;    horseradish peroxidase;   

DOI  :  10.1016/0014-5793(92)81338-M

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Calcium-dependent regulation of tension and ATPase activity in permeabilized porcine ventricular muscle was lost after incubation with 10 mM vanadate. After transfer from vanadate to a vanadate-free, low-Ca²⁺ solution (pCa > 8), the permeabilized muscle produced 84.8% ± 20.1% (± S.D., n=98) of the isometric force elicited by high Ca2²⁺ (pCa ∼4.5 prior to incubation with vanadate. Transfer back to a high Ca²⁺ solution elicited no additional force (83.2% ± 18.7% of control force). SDS-PAGE and immunoblot analysis of fibers and solutions demonstrated substantial extraction (>90%) of Troponin I (TnI). Calcium dependence was restored after incubation with solutions containing either whole cardiac troponin or a combination of TnI and troponin C subunits. This reversible extraction of troponin directly demonstrates the role of TnI in the regulation of striated muscle contractility and permits specific substitution of the native TnI with exogenously supplied protein.

【 授权许可】

Unknown