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In the present study, the carbon monoxide (CO) sensing property of Cu(II)−Laspartic acid nanofibers/polyaniline (PANI) nanofibers composite was investigated at room temperature. The nanofiber composite was formed through the ultrasound mixing of emeraldine salt PANI nanofibers and Cu(II)−L-aspartic acid nanofibers, which were synthesized by using a polymerization process and simple self-assembly method, respectively. The nanofibers composite demonstrated a branched structure in which the Cu(II)−L-aspartic acid nanofiber framework is similar to the trunk of a tree and the polyaniline nanofibers is like its branches. It seems that this special structure and one-dimension/one-dimension interface are suitable for gas adsorption and sensing. The performance of the prepared sensor toward CO gas was investigated at room temperature in a wide concentration range (200−8000 ppm). The experimental results indicate that the incorporation of amino acid-based copper metal− biomolecule framework nanofibers to PANI nanofibers enhances the response value (12.41% to 4000 ppm), yielding good selectivity and acceptable response and recovery characteristics (220 s/240 s) at room temperature. The detection limit of Cu(II)−L-aspartic acid nanofibers/PANI nanofibers sensor for carbon monoxide is obtained at 120 ppm.