Temporal Profiling Establishes a Dynamic S-Palmitoylation Cycle.
Temporal Profiling Establishes a Dynamic S-Palmitoylation Cycle.
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时间分析建立动态的S-膜酰化周期。
DOI：
10.1021/acschembio.8b00157
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发表时间：
2018-06-15
影响因子：
4
通讯作者：
Martin BR
中科院分区：
文献类型：
--
作者：
Won SJ;Martin BR
S-棕榈酰化是膜锚定、适当运输以及数百种完整和外周膜蛋白的正常功能所必需的。先前的生物正交脉冲追踪蛋白质组学分析确定Ras家族GTP酶、极性蛋白和G蛋白为对脱棕榈酰酶抑制敏感的快速循环S-棕榈酰化蛋白，但酶调节的动态S-棕榈酰化的广度在很大程度上仍然是一个谜。在这里，我们提出了一种脉冲生物正交S-棕榈酰化测定的S-棕榈酰化动力学的时间分析。低浓度的十六烷基氟膦酸酯（HDFP）失活APT和ABHD 17家族的脱棕榈酰化酶，这大大增加了炔基脂肪酸标记和分层的S-棕榈酰化的蛋白质成动力学不同的亚组。最令人惊讶的是，HDFP处理不影响稳态S-棕榈酰化水平，尽管抑制了所有验证的脱棕榈酰化酶。APT 1 −/−/APT 2 −/−小鼠大脑的S-棕榈酰化分析同样显示S-棕榈酰化水平没有变化。与羟胺开关方法相比，生物正交炔基脂肪酸仅掺入一小部分动态S-棕榈酰化蛋白质中，这提高了S-棕榈酰化比通常表征的更稳定的可能性。总的来说，破坏脱棕榈酰酶活性增强炔基脂肪酸掺入，但不会大大影响整个蛋白质组的稳态S-棕榈酰化。
S-palmitoylation is required for membrane anchoring, proper trafficking, and the normal function of hundreds of integral and peripheral membrane proteins. Previous bioorthogonal pulse-chase proteomics analyses identified Ras family GTPases, polarity proteins, and G proteins as rapidly cycling S-palmitoylated proteins sensitive to depalmitoylase inhibition, yet the breadth of enzyme regulated dynamic S-palmitoylation largely remains a mystery. Here we present a pulsed bioorthogonal S-palmitoylation assay for temporal analysis of S-palmitoylation dynamics. Low concentration hexadecylfluorophosphonate (HDFP) inactivates the APT and ABHD17 families of depalmitoylases, which dramatically increases alkynyl-fatty acid labeling and stratifies S-palmitoylated proteins into kinetically distinct subgroups. Most surprisingly, HDFP treatment does not affect steady-state S-palmitoylation levels, despite inhibiting all validated depalmitoylating enzymes. S-palmitoylation profiling of APT1−/−/APT2−/− mouse brains similarly show no change in S-palmitoylation levels. In comparison with hydroxylamine-switch methods, bioorthogonal alkynyl fatty acids are only incorporated into a small fraction of dynamic S-palmitoylated proteins, raising the possibility that S-palmitoylation is more stable than generally characterized. Overall, disrupting depalmitoylase activity enhances alkynyl fatty acid incorporation, but does not greatly affect steady state S-palmitoylation across the proteome.
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影响因子：
4
作者：
Won SJ;Davda D;Labby KJ;Hwang SY;Pricer R;Majmudar JD;Armacost KA;Rodriguez LA;Rodriguez CL;Chong FS;Torossian KA;Palakurthi J;Hur ES;Meagher JL;Brooks CL 3rd;Stuckey JA;Martin BR
通讯作者：
Martin BR
影响因子：
14.8
作者：
Dekker, Frank J.;Rocks, Oliver;Waldmann, Herbert
通讯作者：
Waldmann, Herbert
影响因子：
4.8
作者：
Martin, Brent R.;Lambert, Nevin A.
通讯作者：
Lambert, Nevin A.
影响因子：
4.4
作者：
Thinon E;Fernandez JP;Molina H;Hang HC
通讯作者：
Hang HC
影响因子：
30.3
作者：
Jones ML;Collins MO;Goulding D;Choudhary JS;Rayner JC
通讯作者：
Rayner JC