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BMP6 binding to heparin and heparan sulfate is mediated by N-terminal and C-terminal clustered basic residues
Biochimica et Biophysica Acta (BBA) - General Subjects ( IF 3 ) Pub Date : 2020-11-21 , DOI: 10.1016/j.bbagen.2020.129799
Andrea Denardo , Stefano Elli , Stefania Federici , Michela Asperti , Magdalena Gryzik , Paola Ruzzenenti , Fernando Carmona , Paolo Bergese , Annamaria Naggi , Paolo Arosio , Maura Poli

Background

The bone morphogenetic protein 6 (BMP6) is a crucial inducer of hepcidin, the peptide hormone that regulates the iron availability in our body. Hepcidin expression is influenced by hepatic heparan sulfate (HS) and by heparin administration, suggesting BMP6 interaction with heparin/HS. The BMP2/4 subfamily has been deeply characterized to have a N-terminal heparin/HS binding domain (HBD), whose basic residues contact the sulfate groups on heparin and HS. Such detailed characterization is still required for other, structurally different BMPs, including BMP6.

Methods

BMP6 peptides encompassing potential HBDs were analysed on heparin-functionalized plates and microcantilevers, and on membrane HS expressing CHO-K1 cells. Monomeric wild-type BMP6 and mutants were produced, substituting the basic residues with non-charged ones, and their affinity to the heparin-column was measured. The BMP6-heparin interaction was also predicted at atomic level by in silico molecular dynamics.

Results

N-terminal and C-terminal BMP6 peptides showed high heparin affinity in solid-phase assays. The mutation of the two sites (R5L, R6S, R7L and K126N, K127N, R129S) abolished the heparin-binding activity of the recombinant monomeric BMP6. Monomeric BMP6 and peptides specifically bound to membrane HS of CHO-K1 cells through the same domains. Molecular dynamic studies supported the role of the two HBDs, suggesting a cooperative behaviour.

Conclusions

In BMP6, N-terminal (R5, R6, R7) and C-terminal (K126, K127, R129) domains mediate the interaction with heparin and HS.

General significance

This study provides the molecular mechanism supporting the use of heparin to sequester BMP6 and inhibit hepcidin expression, a novel clinical approach for high-hepcidin iron disorders.



中文翻译:

BMP6与肝素和硫酸乙酰肝素的结合是由N端和C端成簇的碱性残基介导的

背景

骨形态发生蛋白6(BMP6)是hepcidin的重要诱导剂,hepcidin是一种调节我们体内铁利用率的肽激素。Hepcidin的表达受肝素硫酸肝素(HS)和肝素给药的影响,表明BMP6与肝素/ HS相互作用。BMP2 / 4亚家族已被深深地表征为具有N端肝素/ HS结合域(HBD),其基本残基与肝素和HS上的硫酸根基团接触。其他结构上不同的BMP(包括BMP6)仍然需要进行这种详细的表征。

方法

在肝素功能化的平板和微悬臂上以及膜HS表达的CHO-K1细胞上分析了涵盖潜在HBD的BMP6肽。产生了野生型BMP6单体和突变体,用不带电荷的残基取代了碱性残基,并测量了它们对肝素柱的亲和力。还通过计算机分子动力学在原子水平上预测了BMP6-肝素相互作用。

结果

N端和C端BMP6肽在固相分析中显示出较高的肝素亲和力。两个位点(R5L,R6S,R7L和K126N,K127N,R129S)的突变消除了重组单体BMP6的肝素结合活性。单体BMP6和多肽通过相同的域与CHO-K1细胞的膜HS特异性结合。分子动力学研究支持了这两个HBD的作用,表明了一种合作行为。

结论

在BMP6中,N端(R5,R6,R7)和C端(K126,K127,R129)域介导与肝素和HS的相互作用。

一般意义

这项研究提供了支持使用肝素隔离BMP6和抑制hepcidin表达的分子机制,这是一种针对高hepcidin铁症的新型临床方法。

更新日期:2020-12-01
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