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. Author manuscript; available in PMC: 2020 Nov 1.
Published in final edited form as: J Mol Cell Cardiol. 2019 Sep 7;136:42–52. doi: 10.1016/j.yjmcc.2019.09.002

Figure 4: Cleavage of C27 peptides from fusion proteins and preservation of mAb TnI epitope in HcTnI-C27 peptide.

Figure 4:

(A) The 15% small pore SDS-PAGE gels show the cleavage of Tx3-SUMO-HcTnI-C27 and Tx3-SUMO-HcTnI-C27-H fusion proteins. Western blot using mAb 3C11 against the metal-binding tag in carrier protein detected the intact fusion proteins and the cleaved carrier, as well as the recombinant SUMO protease that has the metal-binding tag for rapid purification. Western blot using mAb TnI-1 detected the HcTnI-C27 fusion protein and free HcTnI-C27 peptide released by SUMO protease digestion but not the carrier protein. The binding of mAb TnI-1 is lost for the HcTnI-C27-H mutant peptide. (B) The preservation of mAb TnI-1 epitope in wild type but not mutant C27 peptide was more clearly demonstrated by Western blot using chemically synthesized peptides. Amido Black stains of the PVDF membranes prior to blocking and mAb incubation are shown to verify the effective blotting of the small peptides. MW, molecular weight.