Palladium nanoparticles enzyme aggregate (PANEA) were prepared from Candida antarctica B lipase and palladium salt by precipitation and subsequent in situ Pd nanoparticle formation. This heterogeneous catalyst was successfully used for the Suzuki–Miyaura cross-coupling reaction between bromobenzene with different phenylboronic acid derivatives under mild reaction conditions and using low Pd amount. The nanocatalyst exhibited the highest catalytic activity in a mixture of methanol/water (1:1), obtaining good to excellent product yields from the cross-coupling reaction. A variety of functional groups were accepted and the catalyst was recycled 4 times without activity loss.