When an electrical potential is applied across a single nanopores it generates an ionic current that can be used to address molecules with high precision. Notably, nanopore with a size comparable to the analyte of interest can now be used to monitor chemical and enzymatic reactions, and to perform long nucleic acid reads. At the moment nanopores made of proteins have a superior stability and a lower electrical noise, and they are more reproducible than nanopore made from solid-state materials. However, the shape and size of pore-forming pores is generally fixed, which limit their use in addressing targeted molecules. Here, I will describe the efforts in our laboratory to design and characterize new protein nanopore, and their engineering to allow the capture of analyte of proteins. These new class of nanopores allows measuring the conformational heterogeneity of enzymes during catalysis, and have applications in molecular diagnostics and single-molecule protein sequencing.