Forming the Phosphate Layer in Reconstituted Horse Spleen Ferritin and the Role of Phosphate in Promoting Core Surface Redox Reactions

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• 作者：Joseph L. Johnson ; Michelle Cannon ; Richard K. Watt ; Richard B. Frankel ; and Gerald D. Watt
• 刊名：Biochemistry
• 出版年：1999
• 出版时间：May 18, 1999
• 年：1999
• 卷：38
• 期：20
• 页码：6706 - 6713
• 全文大小：92K
• 年卷期：v.38,no.20(May 18, 1999)
• ISSN：1520-4995

文摘

Apo horse spleen ferritin (apo HoSF) was reconstituted to various core sizes (100-3500 Fe³⁺/HoSF) by depositing Fe(OH)₃ within the hollow HoSF interior by air oxidation of Fe²⁺. Fe²⁺ and phosphate(P_i) were then added anaerobically at a 1:4 ratio, and both Fe²⁺ and P_i were incorporated into the HoSFcores. The resulting P_i layer consisted of Fe²⁺ and P_i at about a 1:3 ratio which is strongly attached to thereconstituted ferritin mineral core surface and is stable even after air oxidation of the bound Fe²⁺. Thetotal amount of Fe²⁺ and P_i bound to the iron core surface increases as the core volume increases up toa maximum near 2500 iron atoms, above which the size of the P_i layer decreases with increasing coresize. Mössbauer spectroscopic measurements of the P_i-reconstituted HoSF cores using ⁵⁷Fe²⁺ show that⁵⁷Fe³⁺ is the major species present under anaerobic conditions. This result suggests that the incoming⁵⁷Fe²⁺ undergoes an internal redox reaction to form ⁵⁷Fe³⁺ during the formation of the P_i layer. Additionof bipyridine removes the ⁵⁷Fe³⁺ bound in the P_i layer as [⁵⁷Fe(bipy)₃]²⁺, showing that the bound ⁵⁷Fe²⁺has not undergone irreversible oxidation. This result is related to previous studies showing that ⁵⁷Fe²⁺bound to native core is reversibly oxidized under anaerobic conditions in native holo bacterial and HoSFferritins. Attempts to bury the P_i layer of native or reconstituted HoSF by adding 1000 additional ironatoms were not successful, suggesting that after its formation, the P_i layer "floats" on the developing ironmineral core.