Biophysical Insight into the SARS-CoV2 Spike–ACE2 Interaction and Its Modulation by Hepcidin through a Multifaceted Computational Approach
通过多方面的计算方法，对SARS-CoV-2 Spike-ACE2相互作用，以及这一作用的铁调素调节进行生物物理洞察 https://pubs.acs.org/doi/10.1021/acsomega.2c00154
关于SWARM对接（SWARM docking），论文以Web server SWARM为工具分析、研究蛋白质（包括多肽）间的相互作用情况。Web server SWARM基于SwarmDock算法，支持对蛋白质/多肽间的相互作用进行灵活的对接建模，确定或排除可结合性，模拟、计算、呈现结合细节（如对接姿态、结合部位、结合亲和力等等），并对可结合的蛋白/多肽生成、输出复合物的3D结构图。
研究者在SARS-CoV-2 Spike上选取了三个参照氨基酸残基，以它们的空间位置来测定Spike三聚体整体或某个单体/链的角度/仰角，如图C左上区域所示。三个参照残基分别位于Spike的ACE2结合位点区域、AMR附近、茎区（stalk of spike，应该指spike的跨膜区域，膜指病毒包膜），它们在Spike蛋白氨基酸序列中的序号分别为494、538、1042。SARS-CoV-2 Spike蛋白氨基酸序列的长度为1273或1269。
当Spike三聚体接近开放构象时，AMR周围呈现出一个hepcidin25结合位点，hepcidin25将结合到AMR 的外壁，而非穿透AMR，结合到AMR腔中（The rectified spike chain in the trimeric form of spike protein presents a binding site around the AMR for hepcidin25 when it is near to the open conformation...but hepcidin25 is not successful for penetrating into the AMR; instead of binding into the AMR cavity it binds to the external wall of the AMR.）；
当Spike三聚体转变为开放构象时，hepcidin25与AMR的结合亲和力降低，此时RBM成为hepcidin25的更佳结合位点（It is indicated in Figure 13B that the RBM is becoming a better binding site for hepcidin25 upon transition of trimeric spike to the open state...It is indicated in Figure 13C that the affinity of hepcidin25 is decreased for AMR upon transition of spike to the open state.）；
2、SARS-CoV-2 （等某些冠状病毒）Spike尾部区域与铁调素的氨基酸序列相似性是独特且特异的，虽然硬骨鱼（teleosts）和脊椎动物（vertebrates）体内通常含有许多富含半胱氨酸的蛋白质（或多肽），
Moreover, there are many cysteine-rich protein sequences in teleosts and vertebrates in general, yet this similarity to the hepcidin gene family (not merely a one-off sequence) was unique and specific.
但Blast搜索未能发现与SARS-CoV-2 Spike尾部区域相似的其它蛋白或多肽。
The search that found hepcidin did not reveal a range of similarities with other teleost proteins.
Blast，Basic Local Alignment Search Tool（基于局部比对算法的搜索工具），一种基因/蛋白研究的有力工具，基本功能是在生物数据库中搜索指定的基因（核苷酸）序列（/片断）、氨基酸序列（/片断）的匹配序列，提供匹配列表、序列间的比对细节、各种匹配参数、图形化呈现。Blast由NCBI（美国国家生物技术信息中心）提供，可在线自由使用。
论文称，前四个刺突蛋白（对应四种普通感冒冠状病毒）的基序（motif，特定序列片断）与其余序列的相似性似乎要少（there appears to be less similarity in the motif between the first four spike proteins than the rest of the sequences）。
The peptide models of Covidin were highly similar to Hepcidin (Figure 6). The docking with modeled ferroportin showed biochemical conservancies, and the interactions observed also have significant physiological mimicry of host Hepcidin.
Hepcidin和Covidin均与铁转运蛋白细胞外域的中心孔（central pore of Ferroportin's extracellular domains）强烈结合，Covidin-Ferroportin复合物与Hepcidin-Ferroportin复合物有着相似的交互作用特征和结合空间；
Both Hepcidin and Covidin bind strongly to the central pore from the extracellular space of a host iron transporter “ferroportin” and the resulting complex has similar interaction features and binding space as the natural hormone Hepcidin.
Our proteolysis, protein modeling, peptide docking, and MD simulation experiments strongly support functional biological mimicry of Covidin with the natural host Hepcidin hormone.
由于Covidin模仿Hepcidin，因此它应该通过泛素蛋白酶体途径（via the ubiquitin proteasomal pathway）引起铁转运蛋白（ferroportin）降解，因而单独的拮抗剂不能逆转细胞内铁超载。
As Covidin mimics Hepcidin, it should, therefore, cause ferroportin degradation via the ubiquitin proteasomal pathway, whereby antagonists alone cannot reverse the intracellular iron overload.
（铁转运蛋白降解使血清铁降低，）低血清铁导致红细胞生成减少，这可能加速缺氧，使患者病情迅速恶化。根据最近关于SARS-CoV-2入侵红细胞生成细胞的报道，这种（铁转运蛋白降解引发的）缺氧可能会因此而进一步加剧。
The hypoxia might also be accelerated by reduced erythropoiesis due to low serum iron, rapidly deteriorating a patient's condition, and aggravating COVID-19 morbidity. This hypoxia may be further aggravated based on recent reports of SARS-CoV-2 invading erythropoietic cells.98
高水平的循环铁调素可抑制肠道铁吸收，并通过降解细胞铁转运蛋白（ferroportin）将铁锁定在铁储存和铁回收细胞内。这可以防止（血液）循环中铁水平的升高。因此，炎症诱导的铁调素增加（inflammation-induced hepcidin increment）导致全身性低铁血症（低铁血指低血清铁水平，low serum iron levels）。此外，发现了SARS-CoV-2 Spike蛋白与铁调素之间的结构相似性，据推测，病毒（SARS-CoV-2）Spike蛋白可能具有铁调素模拟作用（hepcidin-mimetic action）并诱导铁转运蛋白阻断（ferroportin blockage）。如果这一推测正确，那么这将进一步促进细胞内铁潴留并加剧全身性低铁血症（systemic hypoferremia）。
High levels of circulatory hepcidin inhibit intestinal iron absorption and lock iron within the iron-storing and iron-recycling cells by degrading the cellular-iron transporter ferroportin. This prevents iron entry and subsequent iron elevation in the circulation [111]. Thus, inflammation-induced hepcidin increment leads to systemic hypoferremia (low serum iron levels) [112]. In addition, structural similarity has been identified between hepcidin and SARS‐CoV‐2 spike protein. It has been hypothesized that the viral spike protein may have a hepcidin-mimetic action and induce ferroportin blockage [81]. If correct, then this can further promote intracellular iron retention and contribute to systemic hypoferremia.
In such cases, hypoferremia may be caused due to reasons other than hepcidin elevation. For example, this could be due to the putative hepcidin-mimetic action of the viral protein [81], which would block ferroportin and thereby reduce iron entry into the circulation leading to hypoferremia. Another reason could be elevated serum ferritin. In addition to elevating serum hepcidin, inflammation elevates ferritin levels, as observed in COVID-19 (Table 1). Ferritin can sequester a large number of iron ions, and the higher affinity of iron to ferritin and lactoferrin than transferrin (although transferrin levels are reduced in COVID-19) may assist in this sequestration process and contribute to hypoferremia.
Ehsani（“论文依据二”的作者）强调了SARS-CoV-2 Spike糖蛋白和铁调素多肽之间氨基酸序列的惊人相似性，这一观察结果为SARS-CoV-2的疫苗设计和生物工程抗体开发提供了思路。SARS-CoV-2的铁调素模拟作用（Hepcidin-mimetic action of SARS-CoV-2）可能会显着增加循环铁蛋白和组织铁蛋白（circulating and tissue ferritin），同时诱导SI（serum iron）缺乏。然而，SARS-CoV-2是否利用细胞膜上的铁转运蛋（ferroportin）作为侵入宿主细胞的另一种Spike结合受体，还需要进一步分析。此外，SARS-CoV-2是否通过模仿铁调素直接降解铁转运蛋白（whether SARS-CoV-2 directly degrades ferroportin through mimicking hepcidin），或通过炎症间接上调铁调素以介导铁代谢障碍，也需要仔细研究。
Ehsani highlighted the striking similarity between the amino acid sequence of the SARS-CoV-2 spike glycoprotein and the hepcidin protein (37). This observation provides ideas for vaccine design and bioengineered antibody development for SARS-CoV-2. Hepcidin-mimetic action of SARS-CoV-2 may markedly increase circulating and tissue ferritin, while inducing SI deficiency (32, 33). However, whether SARS-CoV-2 utilizes ferroportin on the cell membrane as another binding receptor for the spike protein to invade host cells requires further analysis. Moreover, whether SARS-CoV-2 directly degrades ferroportin through mimicking hepcidin or indirectly upregulates hepcidin through inflammation to mediate iron dysmetabolism also needs to be carefully investigated.
Esaki demonstrated that the amino acid sequence of the coronavirus spike protein is identical to hepcidin, a protein that acts as the main systemic regulator of iron metabolism. Therefore, this similarity between hepcidin and coronavirus spike protein can lead to a mimetic effect, suggesting that SARS-CoV-2 can increase serum hepcidin and then ferritin, and cause hyperferritinemic syndrome5.
Therefore, the association between dysfunctional hemoglobinopathy and SARS-CoV-2-related hyperferritinemia may affect the oxygen transport capacity of erythrocytes, thereby leading to hypoxia, while causing tissue damage due to non-transferrin bound iron (NTBI), and subsequently releasing free radicals at the inflammation sites6.
。。。
铁毒性（iron toxicity）在COVID-19病理生理学中的角色与SARS-CoV-2的铁调素模拟效应（the hepcidin mimetic effect of SARS-CoV-2）有关，铁转运蛋白（ferroportin）在胃肠道和网状内皮系统中的内化（internalization），导致SF（Serum ferritin，血清铁蛋白）的可用性受阻，这进而导致贫血（anemia）和高铁蛋白血症（hyperferritinemia），并最终造成细胞铁死亡（ferroptosis）。
The role of iron toxicity in the pathophysiology of COVID-19 is related to the hepcidin mimetic effect of SARS-CoV-2, with consequent internalization of ferroportin, both in the gastrointestinal tract and reticuloendothelial system, thereby causing a blockage in the availability of SF, which leads to anemia and hyperferritinemia, and ultimately ferroptosis5,17.
通过模仿铁调素的作用（Through mimicking the action of hepcidin），SARS-CoV-2过度增加组织内（例如肝脏、脾脏、骨髓和肌肉）铁蛋白（ferritin）的浓度，同时减少SF（Serum ferritin）的可用性，从而减少红细胞的产生。循环红细胞（circulating erythrocytes）的减少导致持续性的全身低氧血症（systemic hypoxemia），并阻碍组织氧合（tissue oxygenation），进而引发损伤性的急性呼吸综合症（acute respiratory syndrome）。
Through mimicking the action of hepcidin, SARS-CoV-2 exaggeratedly increases the concentration of intratissue (e.g., liver, spleen, bone marrow, and muscles)ferritin, while there is a reduction in the availability of SF, and consequently, a reduction in erythrocyte production. This decrease in circulating erythrocytes perpetuates systemic hypoxemia and hinders tissue oxygenation, which is already impaired in patients with acute respiratory syndrome.
自COVID-19大流行开始以来，在患有这种疾病的个体中观察到了许多代谢改变（numerous metabolic alterations）。众所周知，SARS-CoV-2能够模仿铁调素的作用（SARS-CoV-2 can mimic the action of hepcidin），改变细胞内铁代谢（altering intracellular iron metabolism），但关于（SARS-CoV-2）对铁循环（iron cycle）其他途径可能造成的后果，仍然存在认识分歧。
Since the beginning of the COVID-19 pandemic, numerous metabolic alterations have been observed in individuals with this disease. It is known that SARS-CoV-2 can mimic the action of hepcidin, altering intracellular iron metabolism, but gaps remain in the understanding of possible outcomes in other pathways involved in the iron cycle.
。。。
COVID-19导致多种铁循环途径（iron cycle pathways）发生改变，铁和铁蛋白（ferritin）水平是反映感染状态、演变（进展）以及疾病预后的标志物。转铁蛋白受体（transferrin receptor）基因表达增加表明铁内化增加（increased iron internalization），并且，SARS-CoV-2对铁调素作用的模仿（the mimicry of hepcidin action by SARS-CoV-2）减少了经由铁转运蛋白的铁输出（iron export via ferroportin），这可以解释由细胞内铁捕获（intracellular trapping）引起的低铁循环水平（low circulating levels of iron）。
COVID-19 causes changes in several iron cycle pathways, with iron and ferritin levels being markers that reflect the state and evolution of infection, as well as the prognosis of the disease. The increased expression of the transferrin receptor gene suggests increased iron internalization and the mimicry of hepcidin action by SARS-CoV-2, reduces iron export via ferroportin, which would explain the low circulating levels of iron by intracellular trapping.
。。。
此外，由于（SARS-CoV-2）病毒的多重交互攻击，COVID-19会发生一系列生化途径失衡（biochemical pathway imbalances），例如铁代谢障碍（iron dysmetabolism）。据推测，这是因为SARS-CoV-2模仿了铁调素的作用（SARS-CoV-2 mimics the action of hepcidin），铁调素与铁转运蛋白（ferroportin）相互作用，导致该复合物（指ferroportin-hepcidin complex）内化和降解（causing the internalization and degradation of this complex），减少了细胞内铁的输出，进而引发高铁蛋白血症（hyperferritinemia）和铁死亡（ferroptosis）。
Furthermore, due to the multiple interactive levels of viral attack, a set of several biochemical pathway imbalances such as iron dysmetabolism occurs in COVID-19. This is supposedly because SARS-CoV-2 mimics the action of hepcidin [5], which interacts with ferroportin, causing the internalization and degradation of this complex, favoring iron entry into the cell and decreasing its export from the interior of cells [6], hyperferritinemia and ferroptosis develop [5].
。。。
除了增加铁蛋白外，有研究（还）指出了SARS-CoV-2模仿铁调素作用，增加循环和组织铁蛋白的能力（the ability of SARS-CoV-2 to mimic the action of hepcidin, increasing circulating and tissue ferritin）。这导致血清铁缺乏（serum iron deficiency）并降低血红蛋白（lowered hemoglobin）。过量的铁与氧分子相互作用产生活性氧（reactive oxygen），导致不同器官诸如肺、肝、肾和心脏的氧化损伤（oxidative damage），这会导致（细胞）铁死亡（ferroptosis）。在细胞外环境方面（Regarding the extracellular environment），血红蛋白病（hemoglobinopathy）和铁代谢异常（iron dysmetabolism）共同损害红细胞运输O2的能力，导致缺氧。从生理上讲，贫血性缺氧（anemic hypoxia）诱导肺血管收缩（pulmonary vasoconstriction）并增加微血管内纤维蛋白的形成（fibrin formation within this microvasculature）。因此，铁调素和铁转运蛋白的失调化相互作用（the dysregulated interaction of hepcidin and ferroportin）能够通过平滑肌增殖（through smooth muscle proliferation） 导致肺动脉高压（pulmonary artery hypertension），这可能有助于阐明COVID-19患者因呼吸衰竭而死亡的病理机制（which perhaps helps elucidate death from respiratory failure in these COVID-19 patients）。
In addition to increased ferritin, studies point to the ability of SARS-CoV-2 to mimic the action of hepcidin, increasing circulating and tissue ferritin. This induces serum iron deficiency and lowered hemoglobin [5]. Excess iron interacts with molecular oxygen generating reactive oxygen species, which contributes to oxidative damage in different organs, such as the lungs, liver, kidneys and heart, which can lead to ferroptosis [7]. Regarding the extracellular environment, the combination of hemoglobinopathy and iron dysmetabolism impairs the ability of erythrocytes to transport O2, resulting in hypoxia. Physiologically, anemic hypoxia induces pulmonary vasoconstriction and increased fibrin formation within this microvasculature. Thus, the dysregulated interaction of hepcidin and ferroportin can lead to pulmonary artery hypertension through smooth muscle proliferation [5], which perhaps helps elucidate death from respiratory failure in these COVID-19 patients.
。。。
关于铁和铁蛋白，本研究可以说支持了这样的假设：SARS-CoV-2，通过模仿铁调素的作用（by mimicking the action of hepcidin），增加铁蛋白的流通（increases circulation of ferritin），同时诱导血清铁和血红蛋白水平缺乏（inducing serum iron and hemoglobin level deficiency）。在这一动态中，患者会出现贫血状态（state of anemia），即使他们有高铁蛋白血症（hyperferritinemic）。（SARS-CoV-2）病毒（Spike）蛋白对铁调素的模拟作用（The mimetic action of the viral protein on hepcidin）能够导致铁转运蛋白阻断（ferroportin blockade）、细胞内铁捕获（intracellular iron trapping）和低铁血症（hypoferremia）。有证据表明，炎症期间，细胞内铁的积累（the accumulation of intracellular iron）也会增加铁蛋白，进一步增加细胞内铁的浓度，（与之有关的铁）代谢失调可导致早期多细胞铁死亡（early multicellular ferropoptosis）。
Considering the results of the present study, in relation to iron and ferritin, it can be said they support the hypothesis that SARS-CoV-2, by mimicking the action of hepcidin, increases the circulation of ferritin while inducing serum iron and hemoglobin level deficiency. In this dynamic, the patient develops a state of anemia, even though they are hyperferritinemic [5]. The mimetic action of the viral protein on hepcidin can lead to ferroportin blockade causing intracellular iron trapping and hypoferremia. There is evidence that during inflammation, the accumulation of intracellular iron also increases ferritin, further increasing the concentration of intracellular iron, evidencing metabolic dysregulation that can lead to early multicellular ferropoptosis [9].
Hepcidin and ferritin levels as markers of immune cell activation during septic shock, severe COVID-19 and sterile inflammation
铁调素和铁蛋白水平在败血性休克、严重COVID-19和无菌炎症中作为免疫细胞激活的标志物 https://www.frontiersin.org/arti ... u.2023.1110540/full
Hyperferritinemia, Low Circulating Iron and Elevated Hepcidin May Negatively Impact Outcome in COVID-19 Patients: A Pilot Study
高铁蛋白血症、低循环铁和铁调素升高不利于COVID-19患者的预后（治疗结果）：一项探索研究 https://www.mdpi.com/2076-3921/11/7/1364
The relationship between serum erythropoietin, hepcidin, and haptoglobin levels with disease severity and other biochemical values in patients with COVID‐19
COVID-19患者血清促红细胞生成素、铁调素和触珠蛋白水平与疾病严重程度和其他生化值之间的关系 https://onlinelibrary.wiley.com/doi/10.1111/ijlh.13479
Minihepcidins are rationally designed small peptides that mimic hepcidin activity in mice and may be useful for the treatment of iron overload
小铁调素是合理设计的小肽，可模拟小鼠铁调素的活性，可能有助于治疗铁过载 https://www.jci.org/articles/view/57693
Guided by this model, we showed that 7–9 N-terminal amino acids of hepcidin, including a single thiol cysteine, comprised the minimal structure that retained hepcidin activity, as shown by the induction of ferroportin degradation in reporter cells.
Further modifications to increase resistance to proteolysis and oral bioavailability yielded minihepcidins that, after parenteral or oral administration to mice, lowered serum iron levels comparably to those after parenteral native hepcidin. Moreover, liver iron concentrations were lower in mice chronically treated with minihepcidins than those in mice treated with solvent alone. Minihepcidins may be useful for the treatment of iron overload disorders.
These genetic studies prompted the exploration of other approaches to increasing circulating hepcidin in iron-overloaded patients with hepcidin deficiency. Synthesis of full-length hepcidin is relatively inefficient, and the half-life of hepcidin in circulation is short because of rapid renal clearance.36 To overcome these limitations, several molecules have been designed to mimic hepcidin activity or stimulate endogenous hepcidin production.2,37 We developed minihepcidins (MHs), short peptides based on the 7-9 N-terminal amino acid segment of hepcidin. We first showed that this N-terminal segment of hepcidin is sufficient to induce FPN-1 internalization and degradation in vitro.38 We then engineered these peptides to increase their half-life and potency, and demonstrated that administration of MHs to mice mimics the iron-restrictive effect of endogenous hepcidin.
论文“Utilizing hepcidin agonists to treat siderophilic infections”（利用铁调素激动剂治疗嗜铁菌感染）小节还提到：
Multiple other strategies have been developed to mimic hepcidin activity or increase production of endogenous hepcidin. These include synthetic full-length human hepcidin,45 another peptide-based hepcidin mimetic,46 small molecule FPN-1 inhibitors,47 or pharmacologic inhibition of Tmprss6 by in vivo targeting and degradation of Tmprss6 messenger RNA,48,49 all displaying similar beneficial effects in preclinical models of hepcidin deficiency.
