title: Conserved tyrosine in the first transmembrane segment of solute:sodium symporters is involved in Na+-coupled substrate co-transport.
abstract: Solute:sodium symporters (SSSs) transport vital molecules across the plasma membrane of all living organisms. vSGLT, the Na(+)/galactose transporter of Vibrio parahemeolyticus, is the only SSS for which high resolution structural information is available, revealing a LeuT-like fold and a Na(+)-binding site analogous to the Na2 site of LeuT. Whereas the core transmembrane segments (TMs) of SSSs share high structural similarity with other transporters of LeuT-like fold, TM1 does not correspond to any TM in those structural homologs and was only resolved for the backbone atoms in the initial vSGLT structure (Protein Data Bank code 3DH4). To assess the role of TM1 in Na(+)-coupled substrate symport by the SSSs, here we have studied the role of a conserved residue in TM1 by computational modeling in conjunction with radiotracer transport and binding studies. Based on our sequence alignment and much topological data for homologous PutP, the Na(+)/proline transporter, we have simulated a series of vSGLT models with shifted TM1 residue assignments. We show that in two converged vSGLT models that retained the original TM1 backbone conformation, a conserved residue, Tyr-19, is associated with the Na(+) binding interaction network. In silico and in vitro mutagenesis of homologous Tyr-14 in PutP revealed the involvement of this conserved residue in Na(+)-dependent substrate binding and transport. Thus, our combined computational and experimental data provide the first clues about the importance of a conserved residue in TM1, a unique TM in the proteins with LeuT-like fold, in the Na(+)-coupled symport mechanism of SSSs.
authors.affiliation: Department of Physiology and Biophysics, Weill Cornell Medical College, New York, New York 10065.
authors.forename: Sonia
authors.initials: S
authors.lastname: Mazier
authors.affiliation: Center for Molecular Recognition & Department of Psychiatry, Columbia University College of Physicians and Surgeons, New York, New York 10032, and the New York State Psychiatric Institute, Division of Molecular Therapeutics, New York, New York 10032. Electronic address: mq2102@columbia.edu.
authors.forename: Matthias
authors.initials: M
authors.lastname: Quick
authors.affiliation: Department of Physiology and Biophysics, Weill Cornell Medical College, New York, New York 10065; HRH Prince Alwaleed Bin Talal Bin Abdulaziz Alsaud Institute for Computational Biomedicine, Weill Cornell Medical College, New York, New York 10065,. Electronic address: les2007@med.cornell.edu.
authors.forename: Lei
authors.initials: L
authors.lastname: Shi
mesh_terms: D020816:Amino Acid Motifs; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001692:Biological Transport / Q000502:physiology; D030562:Databases, Protein; D005690:Galactose / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D012964:Sodium / Q000737:chemistry* / Q000378:metabolism; D013329:Structure-Activity Relationship; D027981:Symporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014736:Vibrio parahaemolyticus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D027981:Symporters; D012964:Sodium; D005690:Galactose