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Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms
ABSTRACT Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero‐trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that s...
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Published in: | The FASEB journal 2015-06, Vol.29 (6), p.2583-2594 |
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creator | Widdows, Kate L. Panitchob, Nuttanont Crocker, Ian P. Please, Colin P. Hanson, Mark A. Sibley, Colin P. Johnstone, Edward D. Sengers, Bram G. Lewis, Rohan M. Glazier, Jocelyn D. |
description | ABSTRACT
Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero‐trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that system L amino acid exchange transporters are not fully obligatory and/or that amino acids are initially present inside the vesicles. To address this, we combined computational modeling with vesicle transport assays and transporter localization studies to investigate the mechanisms mediating [14C]l‐serine (a system L substrate) transport into human placental microvillous plasma membrane (MVM) vesicles. The carrier model provided a quantitative framework to test the 2 hypotheses that l‐serine transport occurs by either obligate exchange or nonobligate exchange coupled with facilitated transport (mixed transport model). The computational model could only account for experimental [14C]l‐serine uptake data when the transporter was not exclusively in exchange mode, best described by the mixed transport model. MVM vesicle isolates contained endogenous amino acids allowing for potential contribution to zero‐trans uptake. Both L‐type amino acid transporter (LAT) 1 and LAT2 subtypes of system L were distributed to MVM, with L‐serine transport attributed to LAT2. These findings suggest that exchange transporters do not function exclusively as obligate exchangers.—Widdows, K. L., Panitchob, N., Crocker, I. P., Please, C. P., Hanson, M. A., Sibley, C. P., Johnstone, E. D., Sengers, B. G., Lewis, R. M., Glazier, J. D. Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms. FASEB J. 29, 2583‐2594 (2015). www.fasebj.org |
doi_str_mv | 10.1096/fj.14-267773 |
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Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero‐trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that system L amino acid exchange transporters are not fully obligatory and/or that amino acids are initially present inside the vesicles. To address this, we combined computational modeling with vesicle transport assays and transporter localization studies to investigate the mechanisms mediating [14C]l‐serine (a system L substrate) transport into human placental microvillous plasma membrane (MVM) vesicles. The carrier model provided a quantitative framework to test the 2 hypotheses that l‐serine transport occurs by either obligate exchange or nonobligate exchange coupled with facilitated transport (mixed transport model). The computational model could only account for experimental [14C]l‐serine uptake data when the transporter was not exclusively in exchange mode, best described by the mixed transport model. MVM vesicle isolates contained endogenous amino acids allowing for potential contribution to zero‐trans uptake. Both L‐type amino acid transporter (LAT) 1 and LAT2 subtypes of system L were distributed to MVM, with L‐serine transport attributed to LAT2. These findings suggest that exchange transporters do not function exclusively as obligate exchangers.—Widdows, K. L., Panitchob, N., Crocker, I. P., Please, C. P., Hanson, M. A., Sibley, C. P., Johnstone, E. D., Sengers, B. G., Lewis, R. M., Glazier, J. D. Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms. FASEB J. 29, 2583‐2594 (2015). www.fasebj.org</description><identifier>ISSN: 0892-6638</identifier><identifier>EISSN: 1530-6860</identifier><identifier>DOI: 10.1096/fj.14-267773</identifier><identifier>PMID: 25761365</identifier><language>eng</language><publisher>United States: Federation of American Societies for Experimental Biology</publisher><subject>Amino Acid Transport System y+ - metabolism ; Amino Acids - metabolism ; Amino Acids - pharmacokinetics ; antiporters ; Biological Transport ; Blotting, Western ; Carbon Radioisotopes ; Cell Membrane - metabolism ; Computer Simulation ; facilitated transport ; Female ; Fluorescent Antibody Technique ; Fusion Regulatory Protein 1, Light Chains - metabolism ; Humans ; Large Neutral Amino Acid-Transporter 1 - metabolism ; LAT2 (SLC7A8) ; Microvilli - metabolism ; Models, Biological ; overshoot phenomena ; Placenta - cytology ; Placenta - metabolism ; Pregnancy ; Research Communication ; Serine - metabolism ; Serine - pharmacokinetics ; Transport Vesicles - metabolism</subject><ispartof>The FASEB journal, 2015-06, Vol.29 (6), p.2583-2594</ispartof><rights>FASEB</rights><rights>FASEB.</rights><rights>The Author(s) 2015 FASEB</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4663-61db98b18c583035706a5867aff71b0ee34cbc08b5034bcf970650537d0c0f6a3</citedby><cites>FETCH-LOGICAL-c4663-61db98b18c583035706a5867aff71b0ee34cbc08b5034bcf970650537d0c0f6a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,780,784,885,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/25761365$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Widdows, Kate L.</creatorcontrib><creatorcontrib>Panitchob, Nuttanont</creatorcontrib><creatorcontrib>Crocker, Ian P.</creatorcontrib><creatorcontrib>Please, Colin P.</creatorcontrib><creatorcontrib>Hanson, Mark A.</creatorcontrib><creatorcontrib>Sibley, Colin P.</creatorcontrib><creatorcontrib>Johnstone, Edward D.</creatorcontrib><creatorcontrib>Sengers, Bram G.</creatorcontrib><creatorcontrib>Lewis, Rohan M.</creatorcontrib><creatorcontrib>Glazier, Jocelyn D.</creatorcontrib><title>Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms</title><title>The FASEB journal</title><addtitle>FASEB J</addtitle><description>ABSTRACT
Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero‐trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that system L amino acid exchange transporters are not fully obligatory and/or that amino acids are initially present inside the vesicles. To address this, we combined computational modeling with vesicle transport assays and transporter localization studies to investigate the mechanisms mediating [14C]l‐serine (a system L substrate) transport into human placental microvillous plasma membrane (MVM) vesicles. The carrier model provided a quantitative framework to test the 2 hypotheses that l‐serine transport occurs by either obligate exchange or nonobligate exchange coupled with facilitated transport (mixed transport model). The computational model could only account for experimental [14C]l‐serine uptake data when the transporter was not exclusively in exchange mode, best described by the mixed transport model. MVM vesicle isolates contained endogenous amino acids allowing for potential contribution to zero‐trans uptake. Both L‐type amino acid transporter (LAT) 1 and LAT2 subtypes of system L were distributed to MVM, with L‐serine transport attributed to LAT2. These findings suggest that exchange transporters do not function exclusively as obligate exchangers.—Widdows, K. L., Panitchob, N., Crocker, I. P., Please, C. P., Hanson, M. A., Sibley, C. P., Johnstone, E. D., Sengers, B. G., Lewis, R. M., Glazier, J. D. Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms. FASEB J. 29, 2583‐2594 (2015). www.fasebj.org</description><subject>Amino Acid Transport System y+ - metabolism</subject><subject>Amino Acids - metabolism</subject><subject>Amino Acids - pharmacokinetics</subject><subject>antiporters</subject><subject>Biological Transport</subject><subject>Blotting, Western</subject><subject>Carbon Radioisotopes</subject><subject>Cell Membrane - metabolism</subject><subject>Computer Simulation</subject><subject>facilitated transport</subject><subject>Female</subject><subject>Fluorescent Antibody Technique</subject><subject>Fusion Regulatory Protein 1, Light Chains - metabolism</subject><subject>Humans</subject><subject>Large Neutral Amino Acid-Transporter 1 - metabolism</subject><subject>LAT2 (SLC7A8)</subject><subject>Microvilli - metabolism</subject><subject>Models, Biological</subject><subject>overshoot phenomena</subject><subject>Placenta - cytology</subject><subject>Placenta - metabolism</subject><subject>Pregnancy</subject><subject>Research Communication</subject><subject>Serine - metabolism</subject><subject>Serine - pharmacokinetics</subject><subject>Transport Vesicles - metabolism</subject><issn>0892-6638</issn><issn>1530-6860</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>24P</sourceid><recordid>eNqFkU2PFCEQhonRuOPozbPh6MFei6b56IuJbhxds4kH9Uxomu5h0sDY0DvuP_BnyzrjZr3opSioh5cqXoSeEzgn0PLXw-6cNFXNhRD0AVoRRqHiksNDtALZ1hXnVJ6hJyntAIAA4Y_RWc0EJ5SzFfp5GbIdZ51dDDgO2ES_X_LvrZ6wj72dXBjxweUt9tZ3sw4W5xLTPs4Zp7z0ziY822urp4SDPWAXkhu3OZUkR6y9CyUa12P7w2x1GO_f9_b2yCWfnqJHQ1Gwz07rGn3bvP968bG6-vzh8uLtVWWaMknFSd-1siPSMEmBMgFcM8mFHgZBOrCWNqYzIDsGtOnM0BaAAaOiBwMD13SN3hx190vnbW9sKN1Maj87r-cbFbVTf1eC26oxXqum4S0tT67Ry5PAHL8vNmXlXTJ2msrPxCUpIkHyWjSU_R_lkglGGBEFfXVEzRxTmu1w1xEBdeuzGnaKNOroc8Ff3J_iDv5jbAHkETi4yd78U0xtvryrN59Ic9L-BZ-OuFA</recordid><startdate>201506</startdate><enddate>201506</enddate><creator>Widdows, Kate L.</creator><creator>Panitchob, Nuttanont</creator><creator>Crocker, Ian P.</creator><creator>Please, Colin P.</creator><creator>Hanson, Mark A.</creator><creator>Sibley, Colin P.</creator><creator>Johnstone, Edward D.</creator><creator>Sengers, Bram G.</creator><creator>Lewis, Rohan M.</creator><creator>Glazier, Jocelyn D.</creator><general>Federation of American Societies for Experimental Biology</general><scope>24P</scope><scope>WIN</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>7T5</scope><scope>H94</scope><scope>5PM</scope></search><sort><creationdate>201506</creationdate><title>Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms</title><author>Widdows, Kate L. ; Panitchob, Nuttanont ; Crocker, Ian P. ; Please, Colin P. ; Hanson, Mark A. ; Sibley, Colin P. ; Johnstone, Edward D. ; Sengers, Bram G. ; Lewis, Rohan M. ; Glazier, Jocelyn D.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c4663-61db98b18c583035706a5867aff71b0ee34cbc08b5034bcf970650537d0c0f6a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Amino Acid Transport System y+ - metabolism</topic><topic>Amino Acids - metabolism</topic><topic>Amino Acids - pharmacokinetics</topic><topic>antiporters</topic><topic>Biological Transport</topic><topic>Blotting, Western</topic><topic>Carbon Radioisotopes</topic><topic>Cell Membrane - metabolism</topic><topic>Computer Simulation</topic><topic>facilitated transport</topic><topic>Female</topic><topic>Fluorescent Antibody Technique</topic><topic>Fusion Regulatory Protein 1, Light Chains - metabolism</topic><topic>Humans</topic><topic>Large Neutral Amino Acid-Transporter 1 - metabolism</topic><topic>LAT2 (SLC7A8)</topic><topic>Microvilli - metabolism</topic><topic>Models, Biological</topic><topic>overshoot phenomena</topic><topic>Placenta - cytology</topic><topic>Placenta - metabolism</topic><topic>Pregnancy</topic><topic>Research Communication</topic><topic>Serine - metabolism</topic><topic>Serine - pharmacokinetics</topic><topic>Transport Vesicles - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Widdows, Kate L.</creatorcontrib><creatorcontrib>Panitchob, Nuttanont</creatorcontrib><creatorcontrib>Crocker, Ian P.</creatorcontrib><creatorcontrib>Please, Colin P.</creatorcontrib><creatorcontrib>Hanson, Mark A.</creatorcontrib><creatorcontrib>Sibley, Colin P.</creatorcontrib><creatorcontrib>Johnstone, Edward D.</creatorcontrib><creatorcontrib>Sengers, Bram G.</creatorcontrib><creatorcontrib>Lewis, Rohan M.</creatorcontrib><creatorcontrib>Glazier, Jocelyn D.</creatorcontrib><collection>Wiley Online Library Open Access</collection><collection>Wiley Online Library Journals</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>Immunology Abstracts</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>The FASEB journal</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Widdows, Kate L.</au><au>Panitchob, Nuttanont</au><au>Crocker, Ian P.</au><au>Please, Colin P.</au><au>Hanson, Mark A.</au><au>Sibley, Colin P.</au><au>Johnstone, Edward D.</au><au>Sengers, Bram G.</au><au>Lewis, Rohan M.</au><au>Glazier, Jocelyn D.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms</atitle><jtitle>The FASEB journal</jtitle><addtitle>FASEB J</addtitle><date>2015-06</date><risdate>2015</risdate><volume>29</volume><issue>6</issue><spage>2583</spage><epage>2594</epage><pages>2583-2594</pages><issn>0892-6638</issn><eissn>1530-6860</eissn><abstract>ABSTRACT
Uptake of system L amino acid substrates into isolated placental plasma membrane vesicles in the absence of opposing side amino acid (zero‐trans uptake) is incompatible with the concept of obligatory exchange, where influx of amino acid is coupled to efflux. We therefore hypothesized that system L amino acid exchange transporters are not fully obligatory and/or that amino acids are initially present inside the vesicles. To address this, we combined computational modeling with vesicle transport assays and transporter localization studies to investigate the mechanisms mediating [14C]l‐serine (a system L substrate) transport into human placental microvillous plasma membrane (MVM) vesicles. The carrier model provided a quantitative framework to test the 2 hypotheses that l‐serine transport occurs by either obligate exchange or nonobligate exchange coupled with facilitated transport (mixed transport model). The computational model could only account for experimental [14C]l‐serine uptake data when the transporter was not exclusively in exchange mode, best described by the mixed transport model. MVM vesicle isolates contained endogenous amino acids allowing for potential contribution to zero‐trans uptake. Both L‐type amino acid transporter (LAT) 1 and LAT2 subtypes of system L were distributed to MVM, with L‐serine transport attributed to LAT2. These findings suggest that exchange transporters do not function exclusively as obligate exchangers.—Widdows, K. L., Panitchob, N., Crocker, I. P., Please, C. P., Hanson, M. A., Sibley, C. P., Johnstone, E. D., Sengers, B. G., Lewis, R. M., Glazier, J. D. Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms. FASEB J. 29, 2583‐2594 (2015). www.fasebj.org</abstract><cop>United States</cop><pub>Federation of American Societies for Experimental Biology</pub><pmid>25761365</pmid><doi>10.1096/fj.14-267773</doi><tpages>12</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Amino Acid Transport System y+ - metabolism Amino Acids - metabolism Amino Acids - pharmacokinetics antiporters Biological Transport Blotting, Western Carbon Radioisotopes Cell Membrane - metabolism Computer Simulation facilitated transport Female Fluorescent Antibody Technique Fusion Regulatory Protein 1, Light Chains - metabolism Humans Large Neutral Amino Acid-Transporter 1 - metabolism LAT2 (SLC7A8) Microvilli - metabolism Models, Biological overshoot phenomena Placenta - cytology Placenta - metabolism Pregnancy Research Communication Serine - metabolism Serine - pharmacokinetics Transport Vesicles - metabolism |
title | Integration of computational modeling with membrane transport studies reveals new insights into amino acid exchange transport mechanisms |
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