Amino acids in transmembrane helix 1 confer major functional differences between human and mouse orthologs of the polyspecific membrane transporter OCT1

Meyer, Marleen J.; Schreier, Pascale C.F.; Başaran, Mert; Vlasova, Stefaniia; Seitz, Tina; Brockmöller, Jürgen; Zdrazil, Barbara; Tzvetkov, Mladen V.

doi:10.1016/j.jbc.2022.101974

Cited by 9 publications

(6 citation statements)

References 53 publications

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“…An additional interesting aspect of the data reported in this study is the ability to draw conclusions about structure-to-function relationships based on substrate-specific differences in the transport kinetics among OCT1 orthologs. In a recent study, we identified amino acid differences at codon 32 (Phe vs. Leu) and codon 36 (Cys vs. Tyr) that confer the differences in the affinities for trospium and fenoterol, respectively, between human and mouse OCT1 [30]. This was supported by docking fenoterol into the available alphafold structural models for human and mouse OCT1.…”

Section: Discussionmentioning

confidence: 86%

“…Interestingly, the intrinsic clearance of fenoterol of mouse OCT1 is also very similar to that of dog and human OCT1 (88.5 µL × min −1 × mg protein −1 ). As a consequence, effects of OCT1 knockout on the hepatic uptake of fenoterol in mice [38] may closely reflect the effects of OCT1 not just in humans [30] but also in dogs. Compared to human OCT1, dog OCT1 had a 5.5-fold higher affinity but a similar capacity for metformin transport, resulting in a 5.8-fold higher intrinsic clearance (Figure 4, Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…One strategy for revealing the mechanisms of transport and polyspecificity is the analysis of the functional differences between OCT1 orthologs in mammals [28]. Using this strategy for human and mouse OCT1, we identified amino acid differences in transmembrane helix 2 (TMH2) and TMH3 that confer differences in the affinity for metformin [29], and the amino acid differences at codons 32 and 36 confer differences in the affinity for trospium and fenoterol, respectively [30]. The same approach may be applied to the dog orthologs.…”

Section: Introductionmentioning

confidence: 99%

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Cloning and Functional Characterization of Dog OCT1 and OCT2: Another Step in Exploring Species Differences in Organic Cation Transporters

Meyer

Falk

Römer

et al. 2022

IJMS

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OCT1 and OCT2 are polyspecific membrane transporters that are involved in hepatic and renal drug clearance in humans and mice. In this study, we cloned dog OCT1 and OCT2 and compared their function to the human and mouse orthologs. We used liver and kidney RNA to clone dog OCT1 and OCT2. The cloned and the publicly available RNA-Seq sequences differed from the annotated exon-intron structure of OCT1 in the dog genome CanFam3.1. An additional exon between exons 2 and 3 was identified and confirmed by sequencing in six additional dog breeds. Next, dog OCT1 and OCT2 were stably overexpressed in HEK293 cells and the transport kinetics of five drugs were analyzed. We observed strong differences in the transport kinetics between dog and human orthologs. Dog OCT1 transported fenoterol with 12.9-fold higher capacity but 14.3-fold lower affinity (higher KM) than human OCT1. Human OCT1 transported ipratropium with 5.2-fold higher capacity but 8.4-fold lower affinity than dog OCT1. Compared to human OCT2, dog OCT2 showed 10-fold lower transport of fenoterol and butylscopolamine. In conclusion, the functional characterization of dog OCT1 and OCT2 reported here may have implications when using dogs as pre-clinical models as well as for drug therapy in dogs.

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Section: Discussionmentioning

confidence: 86%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Cloning and Functional Characterization of Dog OCT1 and OCT2: Another Step in Exploring Species Differences in Organic Cation Transporters

Meyer

Falk

Römer

et al. 2022

IJMS

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“…Differences in substrate uptake between mouse and human homologs of OCT1 are well known and illustrate potential limitations in animal models of drug-OCT1 interactions 39 , 44 . Fenoterol exhibits higher affinity and lower uptake in human OCT1 compared to mouse OCT1, with studies suggesting that residue 36 (Cys in human and Tyr in mouse) dictates this difference 44 .…”

Section: Resultsmentioning

confidence: 99%

Structural basis of promiscuous substrate transport by Organic Cation Transporter 1

Zeng,

Sobti,

Quinn

et al. 2023

Nat Commun

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Organic Cation Transporter 1 (OCT1) plays a crucial role in hepatic metabolism by mediating the uptake of a range of metabolites and drugs. Genetic variations can alter the efficacy and safety of compounds transported by OCT1, such as those used for cardiovascular, oncological, and psychological indications. Despite its importance in drug pharmacokinetics, the substrate selectivity and underlying structural mechanisms of OCT1 remain poorly understood. Here, we present cryo-EM structures of full-length human OCT1 in the inward-open conformation, both ligand-free and drug-bound, indicating the basis for its broad substrate recognition. Comparison of our structures with those of outward-open OCTs provides molecular insight into the alternating access mechanism of OCTs. We observe that hydrophobic gates stabilize the inward-facing conformation, whereas charge neutralization in the binding pocket facilitates the release of cationic substrates. These findings provide a framework for understanding the structural basis of the promiscuity of drug binding and substrate translocation in OCT1.

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“…Over the past few decades, a wealth of functional studies has uncovered several key features of substrate recognition and drug interaction with hOCT1 and hOCT2 [28][29][30][31][32][33][34][35][36] . However, the structural basis of substrate recognition, transport inhibition, DDI, and the transport mechanism of hOCT1 and hOCT2 remain elusive.…”

Section: Mainmentioning

confidence: 99%

Molecular basis of polyspecific drug binding and transport by OCT1 and OCT2

Suo

Wright

Guterres

et al. 2023

Preprint

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A wide range of endogenous and xenobiotic organic ions require facilitated transport systems to cross the plasma membrane for their disposition. In mammals, organic cation transporter subtypes 1 and 2 (OCT1 and OCT2, also known as SLC22A1 and SLC22A2, respectively) are polyspecific transporters responsible for the uptake and clearance of structurally diverse cationic compounds in the liver and kidneys, respectively. Notably, it is well established that human OCT1 and OCT2 play central roles in the pharmacokinetics, pharmacodynamics, and drug-drug interactions (DDI) of many prescription medications, including metformin. Despite their importance, the basis of polyspecific cationic drug recognition and the alternating access mechanism for OCTs have remained a mystery. Here, we present four cryo-EM structures of apo, substrate-bound, and drug-bound OCT1 and OCT2 in outward-facing and outward-occluded states. Together with functional experiments, in silico docking, and molecular dynamics simulations, these structures uncover general principles of organic cation recognition by OCTs and illuminate unexpected features of the OCT alternating access mechanism. Our findings set the stage for a comprehensive structure-based understanding of OCT-mediated DDI, which will prove critical in the preclinical evaluation of emerging therapeutics.

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Amino acids in transmembrane helix 1 confer major functional differences between human and mouse orthologs of the polyspecific membrane transporter OCT1

Cited by 9 publications

References 53 publications

Cloning and Functional Characterization of Dog OCT1 and OCT2: Another Step in Exploring Species Differences in Organic Cation Transporters

Cloning and Functional Characterization of Dog OCT1 and OCT2: Another Step in Exploring Species Differences in Organic Cation Transporters

Structural basis of promiscuous substrate transport by Organic Cation Transporter 1

Molecular basis of polyspecific drug binding and transport by OCT1 and OCT2

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