Purification and characterization of new phytoferritin from black bean (Phaseolus vulgaris L.) seed

In contrast to animal ferritin, relatively little information is available on phytoferritin. Black bean (Phaseolus vulgaris L.) has been consumed in many countries. In the present study, new ferritin from black bean seed was purified by two consecutive anion exchange and size exclusion chromatography. The apparent molecular mass of the native black bean seed ferritin (BSF) was found to be approximately 560 kDa by native PAGE analysis. N-terminal sequence, MALDI-TOF-MS and MS/MS analyses indicate that BSF and soybean seed ferritin (SSF) share very high identity in amino acid sequence. However, SDS-PAGE result indicates that BSF consists of 26.5 (H-1) and 28.0 kDa (H-2) subunits with a ratio of 2 : 1, while the ratio of these two subunits in SSF is 1 : 1. This result demonstrates that the two proteins have different subunit composition which might affect their activities in iron uptake and release. Indeed, at high iron flux, the initial rate of iron oxidative deposition in apoBSF is larger than that in apoSSF. On the contrary, the iron release from BSF is significantly slower than that from SSF. All these results indicate that phytoferritin might regulate the transit of iron into and out of the protein cavity by changing its subunit composition.