Comparative screening of recombinant antigen thermostability for improved leptospirosis vaccine design

Abstract

Recombinant antigens exhibit targeted protectiveproperties and offer important opportunities in the development of therapeutic technologies. Biophysical and structural methods have become important tools for the rational design and engineering of improved antigen-based vaccines. Vaccines containing Leptospira immunoglobulin-like (Lig) protein-derived antigens are currently the most promising candidates for protective immunity against the globally prevalent bacterial pathogen, Leptospira interrogans; however, vaccine trials using these domains have produced inconsistent results. Here, we compare the thermostability of domains from the main immunogenic regions from major leptospiral antigens, LigA and LigB. By measuring temperature-dependent fluorescence decay of the hydrophobic core tryptophan, 17 individual Lig protein immunoglobulin-like (Ig-like) domains were shown to display a broad range of unfolding temperatures. For a majority of the domains, stability issues begin to occur at physiologically relevant temperatures. A set of chimeric Ig-like domains was used to establish the ability of transplanted domain regions to enhance thermostability. Further insights into the determinants for domain stabilization were explored with nuclear magnetic resonance dynamics and mutational analysis. The current study has yielded a set of thermostable Ig-like domain scaffolds for use in engineering antigen-based vaccines and demonstrates the importance of incorporating thermostability screening as a design parameter.

Challenges and Strategies for Developing Recombinant Vaccines against Leptospirosis: Role of Expression Platforms and Adjuvants in Achieving Protective Efficacy

“…In view of the protocol variability, it is also important to consider the quality of the antigens and formulations produced for each study. Ptak et al (2019) [125] demonstrated that the stability of several Lig protein Ig-like domains is affected by variations in physiological temperatures, highlighting the importance of considering thermostability parameters for vaccine production [125]. Taking subunit vaccines based on Lig proteins as an example, the variability of antigen regions selected for each study (Table 1), which can include poor thermostability, could be another important bias that justifies the several inconsistent results observed so far.…”