Tatur Jana, Hagedoorn Peter-Leon, Overeijnder Marieke L, Hagen Wilfred R
Department of Biotechnology, Delft University of Technology, The Netherlands.
Extremophiles. 2006 Apr;10(2):139-48. doi: 10.1007/s00792-005-0484-x. Epub 2005 Dec 10.
A ferritin from the obligate anaerobe and hyperthermophilic archaeon Pyrococcus furiosus (optimal growth at 100 degrees C) has been cloned and overproduced in Escherichia coli to one-fourth of total cell-free extract protein, and has been purified in one step to homogeneity. The ferritin (PfFtn) is structurally similar to known bacterial and eukaryal ferritins; it is a 24-mer of 20 kDa subunits, which add up to a total Mr 480 kDa. The protein belongs to the non-heme type of ferritins. The 24-mer contains approximately 17 Fe (as isolated), 2,700 Fe (fully loaded), or <1 Fe (apoprotein). Fe-loaded protein exhibits an EPR spectrum characteristic for superparamagnetic core formation. At 25 degrees C V(max) = 25 micromole core Fe(3+) formed per min per mg protein when measured at 315 nm, and the K(0.5) = 5 mM Fe(II). At 0.3 mM Fe(II) activity increases 100-fold from 25 to 85 degrees C. The wild-type ferritin is detected in P. furiosus grown on starch. PfFtn is extremely thermostable; its activity has a half-life of 48 h at 100 degrees C and 85 min at 120 degrees C. No apparent melting temperature was found up to 120 degrees C. The extreme thermostability of PfFtn has potential value for biotechnological applications.
来自专性厌氧菌和嗜热古菌激烈火球菌（在100摄氏度下生长最佳）的一种铁蛋白已被克隆，并在大肠杆菌中过量表达，其含量达到无细胞提取物总蛋白的四分之一，且一步纯化至同质。该铁蛋白（PfFtn）在结构上与已知的细菌和真核铁蛋白相似；它是由20 kDa亚基组成的24聚体，总分子量为480 kDa。该蛋白属于非血红素类型的铁蛋白。24聚体含有约17个铁原子（分离时）、2700个铁原子（完全负载时）或少于1个铁原子（脱铁蛋白）。负载铁的蛋白呈现出超顺磁性核心形成的特征性电子顺磁共振光谱。在25摄氏度下，当在315 nm处测量时，V(max) = 每分钟每毫克蛋白形成25微摩尔核心Fe(3+)，K(0.5) = 5 mM Fe(II)。在0.3 mM Fe(II)时，活性从25摄氏度到85摄氏度增加100倍。在以淀粉为生长底物的激烈火球菌中检测到野生型铁蛋白。PfFtn极其耐热；其活性在100摄氏度下的半衰期为48小时，在120摄氏度下为85分钟。在高达120摄氏度时未发现明显的解链温度。PfFtn的极端耐热性在生物技术应用中具有潜在价值。
