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Despite significant efforts to engineer their heterologous production, recombinant spider silk proteins (spidroins) have yet to replicate the unparalleled combination of high strength and toughness exhibited by natural spider silks, preventing their use in numerous mechanically demanding applications. To overcome this long-standing challenge, we have developed a synthetic biology approach combining standardized DNA part assembly and split intein-mediated ligation to produce recombinant spidroins of previously unobtainable size (556 kDa), containing 192 repeat motifs of the Nephila clavipes dragline spidroin. Fibers spun from our synthetic spidroins are the first to fully replicate the mechanical performance of their natural counterparts by all common metrics, i.e., tensile strength (1.03 ± 0.11 GPa), modulus (13.7 ± 3.0 GPa), extensibility (18 ± 6%), and toughness (114 ± 51 MJ/m³). The developed process …