Myosin light chain kinase steady-state kinetics: comparison of smooth muscle myosin II and nonmuscle myosin IIB as substrates.
Myosin light chain kinase steady-state kinetics: comparison of smooth muscle myosin II and nonmuscle myosin IIB as substrates.
复制标题
肌球蛋白轻链激酶稳态动力学：平滑肌肌球蛋白II和非肌肉肌球蛋白IIB作为底物的比较。
DOI：
10.1002/cbf.3209
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发表时间：
2016-10
影响因子：
3.6
通讯作者：
Cremo, Christine R.
中科院分区：
文献类型：
--
作者：
Alcala, Diego B.;Haldeman, Brian D.;Brizendine, Richard K.;Krenc, Agata K.;Baker, Josh E.;Rock, Ronald S.;Cremo, Christine R.
关键词：
肌球蛋白轻链激酶 (MLCK) 磷酸化肌球蛋白调节轻链 (RLC) 的 S19，这是激活肌球蛋白 ATP 酶活性和收缩所必需的。已知平滑肌对炎症、发育阶段或应激等因素表现出可塑性，从而导致非肌肉和平滑肌亚型的差异表达。在这里，我们比较了不同 MLCK 底物磷酸化的稳态动力学参数：(1) 非肌肉 RLC，(2) 平滑肌 RLC，以及 (3) 非肌肉肌球蛋白 IIB 和 (4) 平滑肌肌球蛋白 II 的重肌球蛋白亚片段。我们发现，与非肌肉肌球蛋白 IIB 底物相比，MLCK 对于两种平滑肌肌球蛋白 II 底物的 kcat 高出约 2 倍，而 Km 值非常相似。肌球蛋白轻链激酶对平滑 RLC 和重 meromyosin 的特异性 (kcat/Km) 分别高出 1.6 倍和 1.5 倍，这表明特异性的差异是由 RLC 序列决定的。在 10 个不相同的 RLC 残基中，我们排除了 7 个可能导致不同 MLCK 动力学的根本原因。发现其余 3 个残基在 RLC 的 N 端一半表面暴露，这与它们在底物识别中的重要性一致。这些数据与之前的缺失/嵌合体研究一致，并显着增加了对 MLCK 肌球蛋白相互作用的理解。
Myosin light chain kinase (MLCK) phosphorylates S19 of the myosin regulatory light chain (RLC), which is required to activate myosin's ATPase activity and contraction. Smooth muscles are known to display plasticity in response to factors such as inflammation, developmental stage, or stress, which lead to differential expression of nonmuscle and smooth muscle isoforms. Here, we compare steady-state kinetics parameters for phosphorylation of different MLCK substrates: (1) nonmuscle RLC, (2) smooth muscle RLC, and heavy meromyosin subfragments of (3) nonmuscle myosin IIB, and (4) smooth muscle myosin II. We show that MLCK has a ~2-fold higher kcat for both smooth muscle myosin II substrates compared with nonmuscle myosin IIB substrates, whereas Km values were very similar. Myosin light chain kinase has a 1.6-fold and 1.5-fold higher specificity (kcat/Km) for smooth versus nonmuscle-free RLC and heavy meromyosin, respectively, suggesting that differences in specificity are dictated by RLC sequences. Of the 10 non-identical RLC residues, we ruled out 7 as possible underlying causes of different MLCK kinetics. The remaining 3 residues were found to be surface exposed in the N-terminal half of the RLC, consistent with their importance in substrate recognition. These data are consistent with prior deletion/chimera studies and significantly add to understanding of MLCK myosin interactions.
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影响因子：
5.6
作者：
Baumann, Bruce A. J.;Taylor, Dianne W.;Taylor, Kenneth A.
通讯作者：
Taylor, Kenneth A.
影响因子：
3.9
作者：
Hong, Feng;Haldeman, Brian D.;Jackson, Del;Carter, Mike;Baker, Jonathan E.;Cremo, Christine R.
通讯作者：
Cremo, Christine R.
影响因子：
2.9
作者：
IKEBE, M;HARTSHORNE, DJ
通讯作者：
HARTSHORNE, DJ
影响因子：
4.8
作者：
Haldeman, Brian D.;Brizendine, Richard K.;Cremo, Christine R.
通讯作者：
Cremo, Christine R.
影响因子：
14.2
作者：
Gao, Li;Grant, Audrey V.;Barnes, Kathleen C.
通讯作者：
Barnes, Kathleen C.