Homo and hetero dimerisation of the human guanylate-binding proteins hGBP-1 and hGBP-5 characterised by affinities and kinetics.

The human guanylate-binding proteins (hGBPs) exhibit diverse antipathogenic and tumour-related functions which make them key players in the innate immune response. The isoforms hGBP-1 to hGBP-5 form homomeric complexes and localise to specific cellular compartments. Upon heteromeric interactions, hGBPs are able to guide each other to their specific compartments. Thus, homo- and heteromeric interactions allow the hGBPs to build a network within the cell which might be important for their diverse biological functions. We characterised homomeric complexes of hGBPs in vitro and presented most recently that nonprenylated hGBP-1 and hGBP-5 form dimers as highest oligomeric species while farnesylated hGBP-1 is able to form polymers. We continued to work on the biochemical characterisation of the heteromeric interactions between hGBPs and present here results for nonprenylated hGBP-1 and hGBP-5. Multiangle light scattering identified the GTP-dependent heteromeric complex as dimer. Also hGBP-5's tumour-associated splice variant (hGBP-5ta) was able to form a hetero dimer with hGBP-1. Intriguingly, both hGBP-5 splice variants were able to induce domain rearrangements within hGBP-1. We further characterised the homo and hetero dimers with Förster resonance energy transfer-based experiments. This allowed us to obtain affinities and kinetics of the homo and hetero dimer formation. Furthermore, we identified that the LG domains of hGBP-1 and hGBP-5 build an interaction site within the hetero dimer. Our in vitro study provides mechanistic insights into the homomeric and heteromeric interactions of hGBP-1 and hGBP-5 and present useful strategies to characterise the hGBP network further.