Interaction of antirenalase antibodies with recombinant human renalases 1 and 2 and their C-terminal regions encoded by the alternative exones.

The interaction of antirenalase antibodies with full-length recombinant human renalases RNLS1 and RNLS2, as well as fragments of these proteins encoded by alternative exons 9 and 10 and expressed as fusion proteins with dihydrofolate reductase (DHFR) in Escherichia coli cells has been investigated. In this study we used custom made polyclonal antibodies to the full-length recombinant RNLS1 (amino acid residues (aa) 1-342), created at our request, as well as commercially available monoclonal antibodies to the renalase fragment (aa - 18-342), specific for the RNLS1 isoform and its C-terminal sequence encoded by exon 9. According to Western blot analysis, the antibodies interacted not only with recombinant RNLS1 and RNLS2 preparations, but also with fusion proteins containing C-terminal sequences specific for these isoforms (DHFR-RNLS-9ex and DHFR-RNLS-10ex). The results obtained indicate that the studied antibodies, in addition to their direct targets, also "recognized" other protein constructs of RNLS1 and RNLS2, which were absent in the immunogen preparations used for antibody generation.