In this study, we investigated the impact of surface roughness on the sensitivity of conventional and nanowire-based surface plasmon resonance (SPR) biosensors. The theoretical research was conducted using rigorous coupled-wave analysis with Gaussian surface profiles of gold films determined by atomic force microscopy. The results suggest that, when surface roughness ranges 1 nm, the sensitivity of a conventional SPR system is not significantly affected regardless of the correlation length. For a nanowire-based SPR biosensor, however, we found that the sensitivity degrades substantially with a decreasing correlation length. Particularly, at a correlation length smaller than 100 nm, random rough surface may induce destructive coupling between excited localized surface plasmons, which can lead to prominent reduction of sensitivity enhancement.