The apparent solubility and bioavailability of hydrophobic compounds, the main problems in the bioremediation process, can be overcome by bacteria producing biosurfactants. Lipopeptide biosurfactants, as high-efficiency biosurfactants with biological activity, can be synthesized by nonribosomal peptide synthetase (NRPS). In this study, two strains, Paenibacillus lautus A (HC_A) and Lysinibacillus fusiformis B (HC_B), which can efficiently produce lipopeptide biosurfactants, were screened. The NRPS gene sequences of these two strains were obtained by whole-genome sequencing. The module of synthetic surfactant NRPS from HC_A and the module of synthetic surfactants NRPS and sfp from HC_B were successfully cloned into plasmids and then expressed in E. coli (namely, E. coli pHC_A-NRPS and E. coli pHC_B-NRPS-sfp, respectively). Finally, stable lipopeptide biosurfactants could be expressed heterologously. Our research shows that the constructed recombinant bacteria E. coli pHC_A-NRPS and E. coli pHC_B-NRPS-sfp can heterologously express the NRPS module (graphical abstract is shown in Fig 1). Remarkably, the lipopeptide biosurfactants produced by the constructed recombinant bacteria of the NRPS gene have good emulsifying activity on tributyrin. Meanwhile, adding SiO2 nanoparticles and surface-modified carbon black can synergistically enhance the emulsifying effect, which has not been reported before. This research may deepen the comprehension of microbial strains that produce surfactants to emulsify environmental pollutants and perform bioremediation.