Characterizing glucokinase variant mechanisms using a multiplexed abundance assay.
Characterizing glucokinase variant mechanisms using a multiplexed abundance assay.
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使用多重丰度测定表征葡萄糖激酶变异机制。
DOI：
10.1101/2023.05.24.542036
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发表时间：
2023
期刊：
影响因子：
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通讯作者：
Hartmann-Petersen,Rasmus
中科院分区：
文献类型：
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作者：
Gersing,Sarah;Schulze,TheaK;Cagiada,Matteo;Stein,Amelie;Roth,FrederickP;Lindorff-Larsen,Kresten;Hartmann-Petersen,Rasmus
背景氨基酸替换可以通过多种方式干扰蛋白质的活性。了解它们的机制基础可以精确地确定残基如何有助于蛋白质功能。在这里，我们的特点的机制在人类葡萄糖激酶（GCK）的变体的影响，建立在我们以前的全面研究GCK变体activity.ResultsUsing酵母生长为基础的测定，我们得分的丰度95%的GCK错义和无义变体。当将丰度分数与我们先前确定的活性分数相结合时，我们发现43%的低活性变体也降低了细胞蛋白质丰度。低丰度变体在大结构域中富集，而小结构域中的残基在丰度方面耐受突变。相反，在小结构域的许多变体扰动GCK构象动力学是必不可少的适当activity.ConclusionsIn这项研究中，我们确定了GCK代谢稳定性和构象动力学的重要残基。这些残基可以靶向调节GCK活性，从而影响葡萄糖稳态。
BackgroundAmino acid substitutions can perturb protein activity in multiple ways. Understanding their mechanistic basis may pinpoint how residues contribute to protein function. Here, we characterize the mechanisms underlying variant effects in human glucokinase (GCK) variants, building on our previous comprehensive study on GCK variant activity.ResultsUsing a yeast growth-based assay, we score the abundance of 95% of GCK missense and nonsense variants. When combining the abundance scores with our previously determined activity scores, we find that 43% of hypoactive variants also decrease cellular protein abundance. The low-abundance variants are enriched in the large domain, while residues in the small domain are tolerant to mutations with respect to abundance. Instead, many variants in the small domain perturb GCK conformational dynamics which are essential for appropriate activity.ConclusionsIn this study, we identify residues important for GCK metabolic stability and conformational dynamics. These residues could be targeted to modulate GCK activity, and thereby affect glucose homeostasis.