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Mutations at Tyrosine 88, Lysine 92 and Tyrosine 470 of Human Dopamine Transporter Result in an Attenuation of HIV-1 Tat-Induced Inhibition of Dopamine Transport

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Abstract

HIV-1 transactivator of transcription (Tat) protein disrupts the dopamine (DA) neurotransmission by inhibiting DA transporter (DAT) function, leading to increased neurocognitive impairment in HIV-1 infected individuals. Through integrated computational modeling and pharmacological studies, we have demonstrated that mutation of tyrosine470 (Y470H) of human DAT (hDAT) attenuates Tat-induced inhibition of DA uptake by changing the transporter conformational transitions. The present study examined the functional influences of other substitutions at tyrosine470 (Y470F and Y470A) and tyrosine88 (Y88F) and lysine92 (K92M), two other relevant residues for Tat binding to hDAT, in Tat-induced inhibitory effects on DA transport. Y88F, K92M and Y470A attenuated Tat-induced inhibition of DA transport, implicating the functional relevance of these residues for Tat binding to hDAT. Compared to wild type hDAT, Y470A and K92M but not Y88F reduced the maximal velocity of [3H]DA uptake without changes in the Km. Y88F and K92M enhanced IC50 values for DA inhibition of [3H]DA uptake and [3H]WIN35,428 binding but decreased IC50 for cocaine and GBR12909 inhibition of [3H]DA uptake, suggesting that these residues are critical for substrate and these inhibitors. Y470F, Y470A, Y88F and K92M attenuated zinc-induced increase of [3H]WIN35,428 binding. Moreover, only Y470A and K92M enhanced DA efflux relative to wild type hDAT, suggesting mutations of these residues differentially modulate transporter conformational transitions. These results demonstrate Tyr88 and Lys92 along with Tyr470 as functional recognition residues in hDAT for Tat-induced inhibition of DA transport and provide mechanistic insights into identifying target residues on the DAT for Tat binding.

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Acknowledgments

This research was supported by grant from the National Institute on Drug Abuse to Jun Zhu (DA035714).

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The authors declare no conflicts of interest.

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Authors and Affiliations

  1. Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, 715 Sumter Street, Columbia, SC, 29208, USA

    Narasimha M. Midde, Pamela M. Quizon, Wei-Lun Sun & Jun Zhu

  2. Molecular Modeling and Biopharmaceutical Center, College of Pharmacy, University of Kentucky, Lexington, KY, 40536, USA

    Yaxia Yuan, Xiaoqin Huang & Chang-Guo Zhan

  3. Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY, 40536, USA

    Yaxia Yuan, Xiaoqin Huang & Chang-Guo Zhan

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  1. Narasimha M. Midde
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  2. Yaxia Yuan
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  4. Wei-Lun Sun
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  7. Jun Zhu
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Correspondence to Jun Zhu.

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Midde, N.M., Yuan, Y., Quizon, P.M. et al. Mutations at Tyrosine 88, Lysine 92 and Tyrosine 470 of Human Dopamine Transporter Result in an Attenuation of HIV-1 Tat-Induced Inhibition of Dopamine Transport. J Neuroimmune Pharmacol 10, 122–135 (2015). https://doi.org/10.1007/s11481-015-9583-3

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