Different Na؉ /Cl ؊ -dependent neurotransmitter transporters of the SLC6a family have been shown to form dimers or oligomers in both intracellular compartments and at the cell surface. In contrast, the glycine transporters (GlyTs) GlyT1 and -2 have been reported to exist as monomers in the plasma membrane based on hydrodynamic and native gel electrophoretic studies. Here, we used cysteine substitution and oxidative crosslinking to show that of GlyT1 and GlyT2 also form dimeric complexes within the plasma membrane. GlyT oligomerization at the cell surface was confirmed for both GlyT1 and GlyT2 by fluorescence resonance energy transfer microscopy. Endoglycosidase treatment and surface biotinylation further revealed that complex-glycosylated GlyTs form dimers located at the cell surface. Furthermore, substitution of tryptophan 469 of GlyT2 by an arginine generated a transporter deficient in dimerization that was retained intracellulary. Based on these results and GlyT structures modeled by using the crystal structure of the bacterial homolog LeuT Aa , as a template, residues located within the extracellular loop 3 and at the beginning of transmembrane domain 6 are proposed to contribute to the dimerization interface of GlyTs.
Objective: Structural characterization of 5-hydroxytryptamine (5-HT)2A receptor in homo sapiens using in silico method.Methods: In silico approach has particularly providing a realistic representation needed to understand the fundamental molecular structure of a serotonin receptor. The structure has been generated using Swiss model, Modeller 9.14, Phyre2, and Geno three-dimensional, which was visualized using PyMol, and validated by Procheck and ERRAT analysis along with the values of different secondary structures mapping to diverse sections of the Ramachandran plot.Results: We compared all different models. Further structural analysis suggested that the structure of 5-HT2A is a monomer with 18 alpha helices, seven beta sheets, and one disulfide bridge. There is no signal peptide region in the protein sequence. The structure contains mostly polar and aromatic amino acid as suggested by using hydropathy plot. However, in both partitioning systems bilayer to water and water to bilayer, there are some hydropathy predicted segments, which are also transmembrane segments. Finally, the pore features, including diameter profile, size, and shape, were determined by porewalker, and the shape of the pore was found to be UDSD.Conclusion: This study suggested that 5-HT2A receptor interaction with its natural ligand serotonin and other inhibitor compounds would further additional information about G protein-coupled receptors. The 5-HT2A receptor could be an important target for therapeutics development.
Objective: In silico approach has particularly drawn attention in providing a realistic representation needed to understand the fundamental molecular structure of a transporter. The importance of folate metabolism and role in the internalization of antifolates in eukaryotes have been studied extensively, but the structural study of folate transporters in Homo sapiens (HFT), Plasmodium falciparum (PFT), and Streptococcus sp. (SFT) is still lacking. This study was conducted to study and compare the structures of prokaryotic and eukaryotic folate transporters.Methods: HFT, PFT, and SFT were queried using blast and sequences were retrieved using National Center for Biotechnology and Information (NCBI) databases. This was superseded by structural and functional prediction of transporters. The structure has been generated using Swiss model which was visualized using PyMol and validated by Procheck and ERRAT analysis along with the values of different secondary structures mapping to diverse sections of the Ramachandran plot. The structural and functional comparison was performed by PROSO, ProFunc, TM Score, Porewalker, TMHMM, and Protscale.Result: All the parameters for structural comparison suggest that H. sapiens folate transporter is 16.67% and 17.72% identical to Plasmodium and Streptococcus whereas Plasmodium is 21.59% identical to Streptococcus. The evaluation of transmembrane helices and hydrophobicity resulted in the presence of 1, 4, and 12 membrane-spanning segments with predicted US, UDUD, and UDS as pore shape in Plasmodium, Streptococcus, and humans. Conclusion:Such folate receptors are the main targets for the specific conveyance of antifolates. The differences found between these species may offer possibilities for the development of new drugs in future.
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