The construction of sorbent with excellent structure and highly dispersed active components is vital to improving its desulfurization activity. Herein, the Fe nanoparticles/carbon nanofibers (Fe/CNFs) desulfurization sorbents were fabricated via electrospinning, template method and microwave carbonization. The structure of sorbents was successfully regulated with the adjustments of Fe content and the types of sacrificial phases. It was found that the Fe content impacted strongly on both structural and desulfurization performance of the sorbents. The sorbents with various Fe contents showed a trend of first increasing and then decreasing with the increased Fe content in breakthrough sulfur capacity. Among the various samples, the sample with the highest sulfur content was 7.21 g S/100 g sorbent. Furthermore, the Fe/CNFs sorbents were optimized via soft and hard template methods, by taking polymethyl methacrylate (PMMA) and SiO2 nanospheres as sacrificial phases. Desulfurization tests implied that the PMMA and SiO2 modified sorbents involved sulfur capacity of 13.76 and 14.18 g S/100 g sorbent, respectively. According to the characterizations, the modified sorbents were imparted with micro- and open-type macro-porous structure, respectively, upon which, the active components were uniformly dispersed and exposed. The excellent performance of sorbent could be attributed to the synergistic effect of porous structure and widely distributed Fe nanoparticles on CNFs. This work highlights new strategy for the synthesis of desulfurization sorbent.