Mutagenic Analysis of the Putative ABCC6 Substrate-Binding Cavity Using a New Homology Model

International Journal of Molecular Sciences

D. Peter Tieleman¹  Valentina Corradi¹  Sylvia Donnelly²  Koen van de Wetering²  Flora Szeri²  Fatemeh Niaziorimi²  Gwenaëlle Conseil³  Susan P. C. Cole³ 

[1] Department of Biological Sciences and Centre for Molecular Simulation, University of Calgary, Calgary, AB T2N 1N4, Canada;Department of Dermatology and Cutaneous Biology and PXE Center of Excellence in Research and Clinical Care, Thomas Jefferson University, Philadelphia, PA 19107, USA;Department of Pathology and Molecular Medicine, Queen’s University, Kingston, ON K7L 3N6, Canada;

关键词: ABC transporter;    pseudoxanthoma elasticum;    homology modeling;    substrate-binding site;    cellular ATP efflux;    mutagenesis;   

DOI  :  10.3390/ijms22136910

来源: DOAJ

【 摘 要 】

Inactivating mutations in ABCC6 underlie the rare hereditary mineralization disorder pseudoxanthoma elasticum. ABCC6 is an ATP-binding cassette (ABC) integral membrane protein that mediates the release of ATP from hepatocytes into the bloodstream. The released ATP is extracellularly converted into pyrophosphate, a key mineralization inhibitor. Although ABCC6 is firmly linked to cellular ATP release, the molecular details of ABCC6-mediated ATP release remain elusive. Most of the currently available data support the hypothesis that ABCC6 is an ATP-dependent ATP efflux pump, an un-precedented function for an ABC transporter. This hypothesis implies the presence of an ATP-binding site in the substrate-binding cavity of ABCC6. We performed an extensive mutagenesis study using a new homology model based on recently published structures of its close homolog, bovine Abcc1, to characterize the substrate-binding cavity of ABCC6. Leukotriene C4 (LTC₄), is a high-affinity substrate of ABCC1. We mutagenized fourteen amino acid residues in the rat ortholog of ABCC6, rAbcc6, that corresponded to the residues in ABCC1 found in the LTC₄ binding cavity. Our functional characterization revealed that most of the amino acids in rAbcc6 corresponding to those found in the LTC₄ binding pocket in bovine Abcc1 are not critical for ATP efflux. We conclude that the putative ATP binding site in the substrate-binding cavity of ABCC6/rAbcc6 is distinct from the bovine Abcc1 LTC₄-binding site.

【 授权许可】

Unknown