Interaction of the neutral amino acid transporter ASCT2 with basic amino acids

Ndaru, Elias; Garibsingh, Rachel‐Ann A.; Zielewicz, Laura; Schlessinger, Avner; Grewer, Christof

doi:10.1042/bcj20190859

Cited by 4 publications

(5 citation statements)

References 55 publications

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“…Later on, the 3D structure of hASCT2 showed the presence of a narrow space, between scaffold and transport domain, that forms a larger space directed to the intracellular side; this feature is peculiar of hASCT2 ( Freidman et al, 2020 ). Furthermore, recent work showed that, even if lysine is not recognized as a substrate of hASCT2, small amino acids, modified to have basic side chains, can be translocated by hASCT2 and rASCT2 ( Ndaru et al, 2020 ). Also in this case, the basic amino acid substrates do not interact with C467.…”

Section: Discussionmentioning

confidence: 99%

The Human SLC1A5 Neutral Amino Acid Transporter Catalyzes a pH-Dependent Glutamate/Glutamine Antiport, as Well

Scalise

Mazza

Pappacoda

et al. 2020

Front. Cell Dev. Biol.

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ASCT2 is a neutral amino acid transporter, which catalyzes a sodium-dependent obligatory antiport among glutamine and other neutral amino acids. The human ASCT2 over-expressed in Pichia pastoris and reconstituted in proteoliposomes has been employed for identifying alternative substrates of the transporter. The experimental data highlighted that hASCT2 also catalyzes a sodium-dependent antiport of glutamate with glutamine. This unconventional antiport shows a preferred sidedness: glutamate is inwardly transported in exchange for glutamine transported in the counter direction. The orientation of the transport protein in proteoliposomes is the same as in the cell membrane; then, the observed sidedness corresponds to the transport of glutamate from the extracellular to the intracellular compartment. The competitive inhibition exerted by glutamate on the glutamine transport together with the docking analysis indicates that the glutamate binding site is the same as that of glutamine. The affinity for glutamate is lower than that for neutral amino acids, while the transport rate is comparable to that measured for the asparagine/glutamine antiport. Differently from the neutral amino acid antiport that is insensitive to pH, the glutamate/glutamine antiport is pH-dependent with optimal activity at acidic pH on the external (extracellular) side. The stimulation of glutamate transport by a pH gradient suggests the occurrence of a proton flux coupled to the glutamate transport. The proton transport has been detected by a spectrofluorometric method. The rate of proton transport correlates well with the rate of glutamate transport indicating a 1:1 stoichiometry H + : glutamate. The glutamate/glutamine antiport is also active in intact HeLa cells. On a physiological point of view, the described antiport could have relevance in some districts in which a glutamate/glutamine cycling is necessary, such as in placenta.

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Section: Discussionmentioning

confidence: 99%

The Human SLC1A5 Neutral Amino Acid Transporter Catalyzes a pH-Dependent Glutamate/Glutamine Antiport, as Well

Scalise

Mazza

Pappacoda

et al. 2020

Front. Cell Dev. Biol.

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“…We have shown previously that leak anion current can be used to characterize ASCT2 inhibitors whereby substrates tend to “activate” or increase the leak anion current whereas inhibitors tend to “block” or reduce the leak anion current. [ 7,8,12 ] We have also shown previously that ASCT2 inhibition is stereospecific 11 . In this work, we designed four diastereomers of hydroxyhomoserine to study the stereospecifity of ASCT2 based on this scaffold, and to test for hydrophilic interaction in the side chain.…”

Section: Discussionmentioning

confidence: 99%

“…6 ASCT2 is highly expressed in several types of cancers such as melanoma, colon, glioblastoma, acute myeloid leukemia, lung cancer, breast cancer whereby it supplies the nutrient-deprived cancer cells with glutamine. 7 Together, these results suggest that ASCT2 is an important pharmacological target in the treatment of these diseases. Previously published ASCT2 inhibitors based on various amino acid scaffolds such as glutamine (GPNA), diaminobutyric acid (AABA) whose scaffold led to the discovery of widely used inhibitor V9302, serine (serine biphenyl-4-carboxylate), diaminopropionic acid and hydroxyproline sulfonic acid esters have relatively low ASCT2 affinity, Figure 2.…”

Section: Introductionmentioning

confidence: 89%

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Alanine serine cysteine transporter (ASCT) substrate binding site properties probed with hydroxyhomoserine esters

Ndaru

Zielewicz

Shi

et al. 2022

J of Physical Organic Chem

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The glutamine transporter ASCT2 is highly overexpressed in cancer cells. Block of glutamine uptake by ASCT2 is a potential strategy to inhibit growth of cancer cells. However, pharmacology of the ASCT2 binding site is not well established. In this work, we report the computational docking to the binding site, and the synthesis of a new class of ASCT2 inhibitors based on the novel L-hydroxyhomoserine scaffold. While these compounds inhibit the ASCT2 leak anion conductance, as expected for competitive inhibitors, they did not block leak conductance in glutamate transporters (EAAT1-3 and EAAT5).They were also ineffective with respect to subtype ASCT1, which has >57% amino acid sequence similarity to ASCT2. Molecular docking studies agree very well with the experimental results and suggest specific polar interactions in the ASCT2 binding site. Our findings add to the repertoire of ASCT2 inhibitors and will aid in further studies of ASCT2 pharmacology.

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“…Misincorporation of BMAA is only one of many mechanisms acknowledged as possibly contributing to triggering sporadic neurodegenerative diseases (Chiu et al 2011;Arif et al 2014) and for examples since 2017 see (Beri et al 2017;Metcalf et al 2017;D'Mello et al 2017;Potjewyd et al 2017;Powers et al 2017;Engskog et al 2017;Michaelson et al 2017;Díaz-Parra et al 2017;Tan et al 2018a, b;Downing 2018, 2019;Albano and Lobner 2018;Main and Rodgers 2018;Laugeray et al 2018;Lepoutre et al 2018;Pierozan et al 2018Pierozan et al , 2020Gerić et al 2019;Cheng et al 2019;Pierozan and Karlsson 2019;Tedeschi et al 2019;Diaz-parga et al 2020;Li et al 2020;Ndaru et al 2020;Vallerga et al 2020). Chernoff et al (2017) presents multiple lines of evidence to refute that L-BMAA is a substrate for protein incorporation and we address these below.…”

Section: Misincorporation Of Bmaa Into Proteinsmentioning

confidence: 99%

Is Exposure to BMAA a Risk Factor for Neurodegenerative Diseases? A Response to a Critical Review of the BMAA Hypothesis

Dunlop

et al. 2021

Neurotox Res

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In a literature survey, Chernoff et al. (2017) dismissed the hypothesis that chronic exposure to β-N-methylamino-L-alanine (BMAA) may be a risk factor for progressive neurodegenerative disease. They question the growing scientific literature that suggests the following: (1) BMAA exposure causes ALS/PDC among the indigenous Chamorro people of Guam; (2) Guamanian ALS/PDC shares clinical and neuropathological features with Alzheimer’s disease, Parkinson’s disease, and ALS; (3) one possible mechanism for protein misfolds is misincorporation of BMAA into proteins as a substitute for L-serine; and (4) chronic exposure to BMAA through diet or environmental exposures to cyanobacterial blooms can cause neurodegenerative disease. We here identify multiple errors in their critique including the following: (1) their review selectively cites the published literature; (2) the authors reported favorably on HILIC methods of BMAA detection while the literature shows significant matrix effects and peak coelution in HILIC that may prevent detection and quantification of BMAA in cyanobacteria; (3) the authors build alternative arguments to the BMAA hypothesis, rather than explain the published literature which, to date, has been unable to refute the BMAA hypothesis; and (4) the authors erroneously attribute methods to incorrect studies, indicative of a failure to carefully consider all relevant publications. The lack of attention to BMAA research begins with the review’s title which incorrectly refers to BMAA as a “non-essential” amino acid. Research regarding chronic exposure to BMAA as a cause of human neurodegenerative diseases is emerging and requires additional resources, validation, and research. Here, we propose strategies for improvement in the execution and reporting of analytical methods and the need for additional and well-executed inter-lab comparisons for BMAA quantitation. We emphasize the need for optimization and validation of analytical methods to ensure that they are fit-for-purpose. Although there remain gaps in the literature, an increasingly large body of data from multiple independent labs using orthogonal methods provides increasing evidence that chronic exposure to BMAA may be a risk factor for neurological illness.

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Interaction of the neutral amino acid transporter ASCT2 with basic amino acids

Cited by 4 publications

References 55 publications

The Human SLC1A5 Neutral Amino Acid Transporter Catalyzes a pH-Dependent Glutamate/Glutamine Antiport, as Well

The Human SLC1A5 Neutral Amino Acid Transporter Catalyzes a pH-Dependent Glutamate/Glutamine Antiport, as Well

Alanine serine cysteine transporter (ASCT) substrate binding site properties probed with hydroxyhomoserine esters

Is Exposure to BMAA a Risk Factor for Neurodegenerative Diseases? A Response to a Critical Review of the BMAA Hypothesis

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