Conducting polymers like polyaniline (PANI) have gained a lot of interest due to their outstanding electrical and optoelectronic properties combined with their low cost and easy synthesis. To further exploit the performance of PANI, carbon-based nanomaterials like graphene, graphene oxide (GO) and their derivatives can be incorporated in a PANI matrix. In this study, hexamethylene diisocyanate-modified GO (HDI-GO) nanosheets with two different functionalization degrees have been used as nanofillers to develop high-performance PANI/HDI-GO nanocomposites via in situ polymerization of aniline in the presence of HDI-GO followed by ultrasonication and solution casting. The influence of the HDI-GO concentration and functionalization degree on the nanocomposite properties has been examined by scanning electron microscopy (SEM), Raman spectroscopy, X-ray diffraction (XRD), thermogravimetric analysis (TGA), tensile tests, zeta potential and four-point probe measurements. SEM analysis demonstrated a homogenous dispersion of the HDI-GO nanosheets that were coated by the matrix particles during the in situ polymerization. Raman spectra revealed the existence of very strong PANI-HDI-GO interactions via π-π stacking, H-bonding, and hydrophobic and electrostatic charge-transfer complexes. A steady enhancement in thermal stability and electrical conductivity was found with increasing nanofiller concentration, the improvements being higher with increasing HDI-GO functionalization level. The nanocomposites showed a very good combination of rigidity, strength, ductility and toughness, and the best equilibrium of properties was attained at 5 wt % HDI-GO. The method developed herein opens up a versatile route to prepare multifunctional graphene-based nanocomposites with conductive polymers for a broad range of applications including flexible electronics and organic solar cells.