Smooth muscle myosin light chain kinase efficiently phosphorylates serine 15 of cardiac myosin regulatory light chain.

Smooth muscle myosin light chain kinase efficiently phosphorylates serine 15 of cardiac myosin regulatory light chain.
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DOI:
10.1016/j.bbrc.2011.11.044
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发表时间:
2011-12-16
影响因子:
3.1
通讯作者:
Ajtai K
Ajtai K
中科院分区:
生物学4区
文献类型:
--
作者:
Josephson MP;Sikkink LA;Penheiter AR;Burghardt TP;Ajtai K

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人心室肌球蛋白调节轻链(MYL 2)的特异性磷酸化修饰S15处的蛋白质。这种修饰影响MYL 2二级结构并调节心脏组织收缩的Ca 2+敏感性。平滑肌肌球蛋白轻链激酶(smMLCK)是一种普遍存在于子宫中的激酶,也存在于包括心肌在内的其他收缩组织中。重组130 kDa(短)smMLCK在体外磷酸化MYL 2中的S15。使用质谱法直接检测S15上的磷酸基团,验证S15的特异性修饰。SmMLCK还特异性磷酸化猪心室肌球蛋白和鸡砂囊平滑肌肌球蛋白中的肌球蛋白调节轻链S15(平滑肌中的S20),但未能磷酸化兔骨骼肌肌球蛋白中的肌球蛋白调节轻链。磷酸化动力学,使用一种新的荧光方法,消除了放射性同位素的使用,表明类似的Michaelis-Menten Vmax和KM的调节轻链S15磷酸化率在MYL 2,猪心室肌球蛋白,和鸡砂囊肌球蛋白。这些数据表明,smMLCK是一种特异性和有效的激酶,在体外磷酸化MYL 2,心脏,平滑肌肌球蛋白。smMLCK是否在心肌调节或对引起疾病的刺激的反应中起作用尚不清楚,但根据其特异性、动力学和组织表达,应将其视为心脏组织中潜在的重要激酶。
Specific phosphorylation of the human ventricular cardiac myosin regulatory light chain (MYL2) modifies the protein at S15. This modification affects MYL2 secondary structure and modulates the Ca2+ sensitivity of contraction in cardiac tissue. Smooth muscle myosin light chain kinase (smMLCK) is a ubiquitous kinase prevalent in uterus and present in other contracting tissues including cardiac muscle. The recombinant 130 kDa (short) smMLCK phosphorylated S15 in MYL2 in vitro. Specific modification of S15 was verified using the direct detection of the phospho group on S15 with mass spectrometry. SmMLCK also specifically phosphorylated myosin regulatory light chain S15 in porcine ventricular myosin and chicken gizzard smooth muscle myosin (S20 in smooth muscle) but failed to phosphorylate the myosin regulatory light chain in rabbit skeletal myosin. Phosphorylation kinetics, measured using a novel fluorescence method eliminating the use of radioactive isotopes, indicates similar Michaelis-Menten Vmax and KM for regulatory light chain S15 phosphorylation rates in MYL2, porcine ventricular myosin, and chicken gizzard myosin. These data demonstrate that smMLCK is a specific and efficient kinase for the in vitro phosphorylation of MYL2, cardiac, and smooth muscle myosin. Whether smMLCK plays a role in cardiac muscle regulation or response to a disease causing stimulus is unclear but it should be considered a potentially significant kinase in cardiac tissue on the basis of its specificity, kinetics, and tissue expression.
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