Glycoproteins with enveloped viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), influenza virus, and human immunodeficiency virus (HIV), display a trimeric conformation. Different from the monomeric form, the trimeric proteins exhibit superior immunogenicity. Several trimerization motifs, such as Foldon derived from phage T4 fibritin, have been used to promote the formation of trimeric proteins with natural conformations. Although the Foldon-induced trimeric proteins are stable, their high immunogenicity limits applications in the development of vaccine antigens. In a previous study, we developed a recombinant human collagen type Ⅲ protein and determined its crystal structure, revealing a triple-helix conformation. However, the potential of this recombinant protein as a trimerization motif remained unknown. In this study, we demonstrated that the recombinant humanized type Ⅲ collagen (Rh3C) was able to act as a trimerization motif, facilitating the spontaneous trimer formation of the Rh3C-conjugated receptor-binding domain (RBD) within the spike (S) protein of SARS-CoV-2. This trimeric protein could induce a stronger SARS-CoV-2 RBD-specific IgG, IgG1, and IgG2a immune response, when compared with the monomeric RBD protein in the immunized mice. Notably, the Rh3C-RBD protein, when adjuvanted with the novel STING agonist CF501, also elicited significantly higher neutralizing antibody responses against both the pseudotyped SARS-CoV-2 (D614G) and its variant Omicron (BA.2.2) in the immunized mice. To showcase the broad applications of the Rh3C trimerization motif, we further demonstrated that the Rh3C-conjugated HA1 of the influenza virus could also elicit a stronger antibody response than free HA1. Considering the wide distribution of the Rh3C protein in human bodies, its use as a trimerization motif would not induce an immune response due to immune tolerance, thereby allowing the immune response to concentrate on targeted viral proteins. Therefore, this Rh3C-based trimerization motif holds great potential for the design and optimization of vaccines consisting of trimeric protein antigens. {L-End}