title: ABC transporter Cdr1p harbors charged residues in the intracellular loop and nucleotide-binding domain critical for protein trafficking and drug resistance.
abstract: The ABC transporter Cdr1 protein of Candida albicans, which plays a major role in antifungal resistance, has two transmembrane domains (TMDs) and two nucleotide-binding domains (NBDs). The 12 transmembrane helices of TMDs that are interconnected by extracellular and intracellular loops (ICLs) mainly harbor substrate recognition sites where drugs bind while cytoplasmic NBDs hydrolyze ATP which powers drug efflux. The coupling of ATP hydrolysis to drug transport requires proper communication between NBDs and TMDs typically accomplished by ICLs. This study examines the role of cytoplasmic ICLs of Cdr1p by rationally predicting the critical residues on the basis of their interatomic distances. Among nine pairs that fall within a proximity of <4 Å, an ion pair between K577 of ICL1 and E315 of NBD1 was found to be critical. The substitution, swapping and changing of the length or charge of K577 or E315 by directed mutagenesis led to a misfolded, non-rescuable protein entrapped in intracellular structures. Furthermore, the equipositional ionic pair-forming residues from ICL3 and NBD2 (R1260 and E1014) did not impact protein trafficking. These results point to a new role for ICL/NBD interacting residues in PDR ABC transporters in protein folding and trafficking.
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
authors.forename: Abdul Haseeb
authors.initials: AH
authors.lastname: Shah
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
authors.forename: Atanu
authors.initials: A
authors.lastname: Banerjee
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
authors.forename: Manpreet Kaur
authors.initials: MK
authors.lastname: Rawal
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
authors.forename: Ajay Kumar
authors.initials: AK
authors.lastname: Saxena
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
authors.forename: Alok Kumar
authors.initials: AK
authors.lastname: Mondal
authors.affiliation: School of Life Sciences, Jawaharlal Nehru University, New Delhi 110067, India rp47jnu@gmail.com.
authors.forename: Rajendra
authors.initials: R
authors.lastname: Prasad
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D002176:Candida albicans / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D026141:Drug Resistance, Multiple, Fungal / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D007522:Isocitrate Lyase / Q000235:genetics; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D009154:Mutation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000235:genetics*
chemical_list: D018528:ATP-Binding Cassette Transporters; C094693:CDR1 protein, Candida albicans; D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D007522:Isocitrate Lyase
keywords: ABC transporter; Candida albicans; Cdr1p; intracellular loops; multidrug resistance; nucleotide-binding domains