Multifunctional fibers represent a cornerstone of human civilization, playing a pivotal role in numerous aspects of societal development. Natural biomaterials, in contrast to synthetic alternatives, offer environmental sustainability, biocompatibility, and biodegradability. Among these biomaterials, natural silk is favored in biomedical applications and smart fiber technology due to its accessibility, superior mechanical properties, diverse functional groups, controllable structure, and exceptional biocompatibility. This review delves into the intricate structure and properties of natural silk fibers and their extensive applications in biomedicine and smart fiber technology. It highlights the critical significance of silk fibers in the development of multifunctional materials, emphasizing their mechanical strength, biocompatibility, and biodegradability. A detailed analysis of the hierarchical structure of silk fibers elucidates how these structural features contribute to their unique properties. The review also encompasses the biomedical applications of silk fibers, including surgical sutures, tissue engineering, and drug delivery systems, along with recent advancements in smart fiber applications such as sensing, optical technologies, and energy storage. The enhancement of functional properties of silk fibers through chemical or physical modifications is discussed, suggesting broader high-end applications. Additionally, the review addresses current challenges and future directions in the application of silk fibers in biomedicine and smart fiber technologies, underscoring silk's potential in driving contemporary technological innovations. The versatility and sustainability of silk fibers position them as pivotal elements in contemporary materials science and technology, fostering the development of next-generation smart materials.