The carbohydrate-binding domain of overexpressed STBD1 is important for its stability and protein-protein interactions.
The carbohydrate-binding domain of overexpressed STBD1 is important for its stability and protein-protein interactions.
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DOI：
10.1042/bsr20140053
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发表时间：
2014-07-01
影响因子：
4
通讯作者：
Cheng A
中科院分区：
文献类型：
--
作者：
Zhu Y;Zhang M;Kelly AR;Cheng A
STBD1(淀粉结合结构域包含蛋白1)属于CBM20(家族20碳水化合物结合模块)蛋白质组，与糖原代谢和自噬有关。然而，人们对其监管或互动伙伴知之甚少。在这里，我们证明了STBD1的CBM20对于它的稳定性和与糖原相关蛋白相互作用的能力是至关重要的。该结构域中保守的色氨酸残基(W293)的突变消除了STBD1与碳水化合物直链淀粉结合的能力。与野生型WT蛋白相比，该突变体表现出快速降解，并在抑制蛋白酶体后被挽救。此外，STBD1在COS细胞中表达时会发生泛素化，这一过程需要N-端。相反，抑制自噬并不显著影响蛋白质的稳定性。在过表达实验中，我们发现STBD1与几种糖原相关蛋白相互作用，如GS(糖原合成酶)、GDE(糖原去分支酶)和Laforin。重要的是，STBD1的W293突变体无法做到这一点，这表明CBM20结构域在蛋白质-蛋白质相互作用中发挥了额外的作用。在HepG2肝癌细胞中，STBD1的过表达可能与内源性GS有关。这种结合在糖原分解过程中增加，表明糖原不是连接这种相互作用所必需的。综上所述，我们的结果揭示了对STBD1的监管和约束合作伙伴的新见解。STBD1是一种含有碳水化合物结合域的蛋白质，与自噬和糖原代谢有关。在这里，我们展示了碳水化合物结合结构域对于它的稳定性和与几种糖原相关蛋白结合的能力是至关重要的。
STBD1 (starch-binding domain-containing protein 1) belongs to the CBM20 (family 20 carbohydrate binding module) group of proteins, and is implicated in glycogen metabolism and autophagy. However, very little is known about its regulation or interacting partners. Here, we show that the CBM20 of STBD1 is crucial for its stability and ability to interact with glycogen-associated proteins. Mutation of a conserved tryptophan residue (W293) in this domain abolished the ability of STBD1 to bind to the carbohydrate amylose. Compared with the WT (wild-type) protein, this mutant exhibited rapid degradation that was rescued upon inhibition of the proteasome. Furthermore, STBD1 undergoes ubiquitination when expressed in COS cells, and requires the N-terminus for this process. In contrast, inhibition of autophagy did not significantly affect protein stability. In overexpression experiments, we discovered that STBD1 interacts with several glycogen-associated proteins, such as GS (glycogen synthase), GDE (glycogen debranching enzyme) and Laforin. Importantly, the W293 mutant of STBD1 was unable to do so, suggesting an additional role for the CBM20 domain in protein–protein interactions. In HepG2 hepatoma cells, overexpressed STBD1 could associate with endogenous GS. This binding increased during glycogenolysis, suggesting that glycogen is not required to bridge this interaction. Taken together, our results have uncovered new insights into the regulation and binding partners of STBD1. STBD1 is a protein with a carbohydrate-binding domain that is implicated in autophagy and glycogen metabolism. Here we show the carbohydrate-binding domain is crucial for its stability and ability to bind to several glycogen-associated proteins.