More about the Zinc Link between Ceruloplasmin and Transferrin: The Distinct Roles of Copper and Zinc for Moving Iron

by Jon Sasmor RCPC (Mineral Guide, MinBalance LLC)
Updated March 4, 2022

This article provides further answers from the researchers and further reflections about the Zinc Link. If you haven't read it already, you probably want to start with the previous article, New! The Zinc Link: Zinc's Separate, Critical Role in Copper-Iron Metabolism, Linking Ceruloplasmin with Transferrin.

According to the research of Sakajiri et al. (2021), zinc links ceruloplasmin with apo-transferrin to allow iron(III) to pass safely from ceruloplasmin to apo-transferrin.

Further Answers from the Zinc Link Researchers

Here are additional answers kindly provided by Dr. Takaki Yamamura, corresponding author, reprinted gratefully by permission:

  1. If ferroportin, holo-ceruloplasmin, and apo-transferrin all are present, but zinc is missing, does all of the iron(III) eventually leak into the blood as non-transferrin bound iron(III) [NTBI]? Or, does some iron(III) get stuck or stored in ceruloplasmin? Does some iron get returned back into the cell as iron(II)?

    • I think that about 60 % of Fe(III) ions on ceruloplasmin are captured by apo-transferrin, some are leaked into the blood as NTBI, and the rest (if any) are unknown (Fig. 2 of our paper).

  2. If ferroportin and apo-transferrin are present, but holo-ceruloplasmin is missing, does iron(II) leak from the cell and auto-oxidize, or does iron(II) all remain in the cell?

    • As described in the Introduction of our paper, “Importance of ceruloplasmin (CP) in vivo is demonstrated in aceruloplasminemia, a genetic disease caused by mutations in the CP gene, resulting in CP protein deficiency and subsequent iron deposition mainly in the brain, liver, and pancreas. Iron deposition in the brain leads to neurodegenerative diseases. A study showed that CP-knockout (CP-/-) mice had increased iron deposition, which led to free radical injury in the central nervous system.” See the References 13-16 of our paper.

  3. The article states "The safe transfer of iron from FPN1 to apo-TF via MCF would require the formation of a ternary complex of FPN1, MCF, and apo-TF. If zinc-bound MCF can bind to FPN1, it may allow the formation of the ternary complex. Obtaining evidence for the presence of a ternary complex of FPN1, zinc-bound MCF, and apo-TF is a topic for future research."

    Why would ferroportin, zinc-bound-ceruloplasmin, and apo-transferrin form a ternary complex, rather than a sequential pairwise interaction: first ferroportin with zinc-bound ceruloplasmin, and then zinc-bound ceruloplasmin with apo-transferrin?

    • As you said, I think it’s perfectly fine that ferroportin first binds with zinc-bound MCF, and then zinc-bound MCF binds with apo-transferrin. There are already papers that　ferroportin and MCF combine. The question is whether ferroportin and zinc-bound MCF bind. See the References 55 and 56.

  4. Which other metals were excluded as less effective than zinc in mediating the ceruloplasmin-transferrin interaction?

    • In the surface plasmon resonance (SPR) measurement, it is known that when the binding of a small analyte such as a metal to the protein immobilized on a sensor chip in SPR induces a conformational change, an increase in RU (response unit) is observed. Indeed, the injection of Zn(II) into the flow cell in which holo-ceruloplasmin was immobilized on the sensor chip showed a concentration-dependent increase in RU, indicating that the binding of Zn(II) to ceruloplasmin induced a conformational change in the protein. Additionally, we examined the binding of other metal ions. The result was as follows: Zn(II) >> Ni(II) > Co(II) > Mg(II) ≒ Al(III) ≒ Cu(II) >> Ca(II) in order of RU value. RU values of all examined metal ions were extremely small. Therefore, the conformational change is thought to be very small. It is not known how this conformational change contributes to the binding between ceruloplasmin and apo-transferrin, but at least it seems to reflect the difference in affinity between these metal ions and ceruloplasmin. Some similar experiments were performed on copper ions as on zinc ions, but copper ions were not as effective as zinc ions on the transfer of Fe (III) from CP to apo-TF.

  5. Does zinc also mediate an interaction between apo-transferrin and hephaestin?

    • We haven't experimented with hephaestin yet.
      Therefore, we do not currently know if zinc mediates the interaction between hephaestin and apo-TF.

Implications: Significance of Copper and Zinc to Ceruloplasmin Function

The Role of Copper

Copper is needed to activate ceruloplasmin. Ceruloplasmin, in turn, is needed to oxidize iron from +2 to +3 oxidation state, so that iron may safely be transported through the blood by transferrin.

Without copper-loaded ceruloplasmin, severe iron deposition occurs, as iron becomes stuck without egress from cells. Iron accumulates in the brain, liver, pancreas, retina, and other organs. Without ceruloplasmin, these organs show decay by around the fourth or fifth decade of life. (See Dr. Yamamura's answer #2 above; Harris, et al., 1995; Hellman & Gitlin, 2002; Patel, et al., 2002; Yoshida, et al., 1995.)

This condition, when occurring genetically, is called aceruloplasminemia. In the homozygous form (from both parents) of aceruloplasminemia, serum ceruloplasmin and serum copper fall to near zero, serum iron falls by half, and serum ferritin rises sharply. (Yoshida, et al., 1995.) Anemia may develop, which will be worsened by adding iron and corrected by adding ceruloplasmin (Harris, et al., 1995).

A similar pattern involving partial loss of ceruloplasmin function may begin to occur in many of us in states of high stress or copper deficiency. This is why we follow the Root Cause Protocol to restore ceruloplasmin's copper-based oxidase function.

We crucially need copper to allow ceruloplasmin to oxidize iron. Then iron remains properly in action moving oxygen, instead of stuck in tissues causing damaging oxidative stress.

The Role of Zinc

Zinc plays a specific, important role in ceruloplasmin's function, recently discovered by Sakajiri, et al. (2021). Zinc forms a link between ceruloplasmin and apo-transferrin, so that iron(III) may be safely transferred.

Without zinc, transfer of iron(III) from ceruloplasmin to apo-transferrin becomes less effective by up to 40%. Also, without zinc, some iron leaks into the bloodstream as harmful reactive non-transferrin-bound iron (NTBI). (See Dr. Yamamura's answer #1 above).

Zinc cannot replace copper in facilitating ceruloplasmin's ferroxidase function. Nor can copper (or other metals listed in Dr. Yamamura's answer #4 above) replace zinc in mediating the link between ceruloplasmin and apo-transferrin.

As Sakajiri et al. (2021) note, further research analyzing clinical data may reveal the correlation of reduced zinc availability in the blood with increased leaky release of NTBI. NTBI blood testing is available in research laboratories and may become more widely available in the future.

Of course, copper plays many wonderful roles in ceruloplasmin's functions. With resourcefulness and precision, Sakajiri et al. (2021) have honed on in a specific, important additional role for zinc in the transport and safekeeping of iron.

Please see my previous post about the Zinc Link for more information about organic ancestral whole food zinc sources and more details about the Sakajiri et al. (2021) research.

References

Sakajiri, T., Nakatsuji, M., Teraoka, Y., Furuta, K., Ikuta, K., Shibusa, K., Sugano, E., Tomita, H., Inui, T., & Yamamura, T. (2021). Zinc mediates the interaction between ceruloplasmin and apo-transferrin for the efficient transfer of Fe (III) ions. Metallomics, 13(12), mfab065. https://doi.org/10.1093/mtomcs/mfab065

The following references are cited in Sakajiri et al. (2021) and mentioned above in Dr. Yamamura's answers:

Reference 13:
Hellman, N. E., & Gitlin, J. D. (2002). Ceruloplasmin metabolism and function. Annual Review of Nutrition, 22(1), 439-458. https://doi.org/10.1146/annurev.nutr.22.012502.114457

Reference 14:
Harris, Z. L., Takahashi, Y., Miyajima, H., Serizawa, M., MacGillivray, R. T., & Gitlin, J. D. (1995). Aceruloplasminemia: molecular characterization of this disorder of iron metabolism. Proceedings of the National Academy of Sciences, 92(7), 2539-2543. https://doi.org/10.1073/pnas.92.7.2539

Reference 15:
Yoshida, K., Furihata, K., Takeda, S., Nakamura, A., Yamamoto, K., Morita, H., Himayuta, S., Ikeda, S., Shimizu, N., & Yanagisawa, N. (1995). A mutation in the ceruloplasmin gene is associated with systemic hemosiderosis in humans. Nature Genetics, 9(3), 267-272. https://doi.org/10.1038/ng0395-267

Reference 16:
Patel, B. N., Dunn, R. J., Jeong, S. Y., Zhu, Q., Julien, J. P., & David, S. (2002). Ceruloplasmin regulates iron levels in the CNS and prevents free radical injury. Journal of Neuroscience, 22(15), 6578-6586. https://doi.org/10.1523/JNEUROSCI.22-15-06578.2002

Reference 55:
Yeh, K. Y., Yeh, M., Mims, L., & Glass, J. (2009). Iron feeding induces ferroportin 1 and hephaestin migration and interaction in rat duodenal epithelium. American Journal of Physiology-Gastrointestinal and Liver Physiology, 296(1), G55-G65. https://doi.org/10.1152/ajpgi.90298.2008

Reference 56:
Jeong, S. Y., & David, S. (2003). Glycosylphosphatidylinositol-anchored ceruloplasmin is required for iron efflux from cells in the central nervous system. Journal of Biological Chemistry, 278(29), 27144-27148. https://doi.org/10.1074/jbc.M301988200