In vitro ubiquitination of creatine kinase B (CKB) and ubiquitous mitochondrial CK (uMtCK) by human ankyrin repeat and suppressor of cytokine signalling (SOCS) box protein 9 (ASB9). Whole cell lysates were prepared from human embryonic kidney 293 (HEK293) cells that stably express null (empty vector), Myc-tagged ASB9 or ASB9ΔSOCS and then immunoprecipitated with anti-CKB antibody (a) or anti-uMtCK antibody (b). The immunocomplexes were analysed by means of in vitro ubiquitination assay with or without ubiquitin. The reaction mixtures were then separated by SDS-PAGE followed by immunoblotting analysis with anti-ubiquitin, anti-CKB, or anti-Myc antibodies. An immunoglobulin heavy chain is shown as an antibody amount control.