Journal
FEBS OPEN BIO
Volume 11, Issue 1, Pages 26-34Publisher
WILEY
DOI: 10.1002/2211-5463.13039
Keywords
ferroportin; hepcidin regulation; iron metabolism; thermostabilization
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Funding
- National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) [R01 DK107309]
- National Health and Medical Research Council (NHMRC) [APP1083536]
- NIDDK [R01 DK082717, P30 DK078392]
- STRIDE Fellowship of the American Physiological Society
- PHS grant from the National Heart, Lung, and Blood Institute (NHLBI) [HL115473]
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This study presents an expression and purification protocol for mouse Fpn, enabling the acquisition of pure protein for further biophysical studies, and paving the way for a better understanding of the transport mechanism of Fpn.
Ferroportin (Fpn) is an essential mammalian iron transporter that is negatively regulated by the hormone hepcidin. Our current molecular understanding of Fpn-mediated iron efflux and regulation is limited due to a lack of biochemical, biophysical and high-resolution structural studies. A critical step towards understanding the transport mechanism of Fpn is to obtain sufficient quantities of pure and stable protein for downstream studies. As such, we detail here an expression and purification protocol for mouse Fpn yielding milligram quantities of pure protein. We have generated deletion constructs exhibiting enhanced thermal stability and which retained iron-transport activity and hepcidin responsiveness, providing a platform for further biophysical studies of Fpn.
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