Calpain-7 binds to CHMP1B at its second α-helical region and forms a ternary complex with IST1
| Autor: | Masatoshi Maki, Hideki Shibata, Emi Goto, Yuki Maemoto, Yohei Osako, Eri Nozawa |
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| Rok vydání: | 2011 |
| Předmět: |
Oncogene Proteins
Protein family Endosomal Sorting Complexes Required for Transport Endosome Calpain Protein Conformation HEK 293 cells General Medicine Biology Biochemistry ESCRT Recombinant Proteins Cell biology HEK293 Cells Microtubule Organelle Humans Cell fractionation Molecular Biology Ternary complex Protein Binding |
| Zdroj: | Journal of biochemistry. 150(4) |
| ISSN: | 1756-2651 |
| Popis: | Some intracellular proteins involved in the endosomal sorting complex required for transport (ESCRT) system have microtubule interacting and transport (MIT) domains and bind to ESCRT-III protein family members named charged multivesicular body proteins (CHMPs) at their C-terminal regions containing MIT-interacting motifs (MIMs). While two types of MIMs (MIM1 and MIM2) have been reported, CHMP1B has MIM1 and IST1 has both MIM1 and MIM2. Previously, we demonstrated that CHMP1B and IST1 directly interacted with a tandem repeat of MIT domains of calpain-7 (CL7MIT) and that autolytic activity of calpain-7 was enhanced by IST1 in vitro but not by overexpression of IST1 in HEK293T cells. In this study, we detected enhancement of autolysis of mGFP-fused calpain-7 by coexpression with CHMP1B and observed further activation by additional coexpression of IST1 in HEK293T cells. We found that CL7MIT interacted with the second α-helical region of CHMP1B but not with the canonical C-terminal region containing MIM1 in vitro. Co-immunoprecipitation assays demonstrated that the interaction between CL7MIT and CHMP1B and between CL7MIT and IST1 became stronger when IST1 or CHMP1B was additionally coexpressed, suggesting formation of ternary complex of calpain-7, IST1 and CHMP1B. Moreover, subcellular fractionation analyses revealed increase of calpain-7 in membrane/organelle fractions by concomitant overexpression of these ESCRT-III family member proteins. |
| Databáze: | OpenAIRE |
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