[Objective] To study the mutation phenotype of the methyl-accepting chemotaxis protein Tlp1 (transducer-like protein) located in the upstream of the gene cluster of Azorhizobium caulinodans ORS571, and to explore its functional mechanism. [Methods] The tlp1 mutant strain was constructed by the homologous recombination and triparental conjugation. Growth of the wild-type strain ORS571 and the tlp1 mutant was compared by measuring OD600 over time of cultures in TY medium. The semi-solid plate assay was used to test the ability of chemotaxis. The production of extracellular polysaccharide and secondary metabolites was measured through Congo red solid culture medium. The acetylene reduction assay was used to measure nitrogen fixation ability. [Results] Compared with the wild type, the growth of the mutant in rich media was the same as that of the wild type. The tlp1 mutant was impaired in chemotaxis to glycerol, and the complemented strain’s chemotactic capability could be partially complemented. The production of extracellular polysaccharides secreted from tlp1 mutant and wild type has no difference, but one of the secondary metabolites — the melanin tlp1 mutant appeared earlier than the wild type. The nitrogenase activity of tlp1 mutant strain was weaker than that of the wild type, and the complemented strain could complement part of the nitrogen fixation ability. [Conclusion] The methyl-accepting chemotaxis protein Tlp1 of A. caulinodans ORS571 showed a certain ability of chemotaxis to glycerol, and also affected the secondary metabolites and the nitrogenase activity in ORS571.