Direct Cloning of Bacterial Surface Polysaccharide Gene Cluster for One-Step Production of Glycoconjugate Vaccine.

Bacterial pathogen infections are fast-growing public health threats and worldwide problems. Glycoconjugate vaccines are among the most effective means in combating such infections. Recent advances in bacterial protein glycan coupling technology (PGCT) have revolutionized the production of glycoconjugate vaccines and drawn enormous attention from both researchers and pharmaceutical companies. Cloning of bacterial surface polysaccharide gene cluster is a prerequisite for the application of PGCT. In this study, we applied the RecET direct cloning strategy for rapid and efficient cloning of O-antigen polysaccharide gene clusters from Escherichia coli serotypes O25b, O26, and O55 in a high-fidelity manner. Then, these gene clusters were applied in PGCT to produce corresponding glycoconjugates. Subsequent immunological studies verified the abilities of glycoconjugate vaccine candidates O25-maltose-binding protein (MBP), O26-MBP, and O55-MBP to generate serotype-specific antibodies and confer protection against E. coli infections. The combination of RecET direct cloning and PGCT makes the rapid production of glycoconjugate vaccines against fast-expanding bacterial pathogens possible.