Superzoom. Italia y el compromiso radical

Italy: the new domestic landscape, la ambiciosa exposición que en 1972 tuvo lugar en el MoMA, supone la consagración y a la vez escenifica la muerte de lo que algunos críticos acababan de catalogar como Arquitectura Radical, término con el que intentaban unificar el trabajo de un heterogéneo y atomizado grupo de arquitectos, principalmente italianos, comprometidos con el replanteamiento total de la definición y objetivos de la disciplina. Se ponía fin así a un corto pero intenso periodo dinamitado por multitud de propuestas experimentales durante el que se generó la última arquitectura con aspiraciones socio-políticas, responsable de un vibrante legado teórico que proclamaba la vuelta a las raíces olvidadas de la profesión y que se mostraba más preocupada por reflexionar sobre sus fundamentos que por perpetuar la práctica de un oficio puesto en crisis. La simbiosis entre el trabajo de Superstudio y Archizoom, dos de los grupos radicales más activos y cuya indisolubilidad hemos querido subrayar acuñando el término ‘superzoom’, articula esta breve revisión a modo de flashback del fenómeno radical italiano, capaz de demostrar en menos de diez años que el ejercicio de la arquitectura y su aplicación docente podían ser ética y políticamente relevantes, abriendo una senda crítica que, en el momento actual de nuestra contradictoria profesión, sería recomendable volver a transitar.

The role of transmembrane domain 2 in cation transport by the Na–K–Cl cotransporter

The human and shark Na–K–Cl cotransporters (NKCC), although 74% identical in amino acid sequence, exhibit marked differences in ion transport and bumetanide binding. We have utilized shark–human chimeras of NKCC1 to search for regions that confer the kinetic differences. Two chimeras (hs3.1 and its reverse sh3.1) with a junction point located at the beginning of the third transmembrane domain were examined after stable transfection in HEK-293 cells. Each carried out bumetanide-sensitive 86Rb influx with cation affinities intermediate between shark and human cotransporters. In conjunction with the previous finding that the N and C termini are not responsible for differences in ion transport, the current observations identify the second transmembrane domain as playing an important role. Site-specific mutagenesis of two pairs of residues in this domain revealed that one pair is indeed involved in the difference in Na affinity, and a second pair is involved in the difference in Rb affinity. Substitution of the same residues with corresponding residues from NKCC2 or the Na-Cl cotransporter resulted in cation affinity changes, consistent with the hypothesis that alternative splicing of transmembrane domain 2 endows different versions of NKCC2 with unique kinetic behaviors. None of the changes in transmembrane domain 2 was found to substantially affect Km(Cl), demonstrating that the affinity difference for Cl is specified by the region beyond predicted transmembrane domain 3. Finally, unlike Cl, bumetanide binding was strongly affected by shark–human replacement of transmembrane domain 2, indicating that the bumetanide-binding site is not the same as the Cl-binding site.

From ab initio quantum mechanics to molecular neurobiology: A cation–π binding site in the nicotinic receptor

The nicotinic acetylcholine receptor is the prototype ligand-gated ion channel. A number of aromatic amino acids have been identified as contributing to the agonist binding site, suggesting that cation–π interactions may be involved in binding the quaternary ammonium group of the agonist, acetylcholine. Here we show a compelling correlation between: (i) ab initio quantum mechanical predictions of cation–π binding abilities and (ii) EC50 values for acetylcholine at the receptor for a series of tryptophan derivatives that were incorporated into the receptor by using the in vivo nonsense-suppression method for unnatural amino acid incorporation. Such a correlation is seen at one, and only one, of the aromatic residues—tryptophan-149 of the α subunit. This finding indicates that, on binding, the cationic, quaternary ammonium group of acetylcholine makes van der Waals contact with the indole side chain of α tryptophan-149, providing the most precise structural information to date on this receptor. Consistent with this model, a tethered quaternary ammonium group emanating from position α149 produces a constitutively active receptor.

Ionophores and receptors using cation-π interactions: Collarenes

Cation-π interactions are important forces in molecular recognition by biological receptors, enzyme catalysis, and crystal engineering. We have harnessed these interactions in designing molecular systems with circular arrangement of benzene units that are capable of acting as ionophores and models for biological receptors. [n]Collarenes are promising candidates with high selectivity for a specific cation, depending on n, because of their structural rigidity and well-defined cavity size. The interaction energies of [n]collarenes with cations have been evaluated by using ab initio calculations. The selectivity of these [n]collarenes in aqueous solution was revealed by using statistical perturbation theory in conjunction with Monte Carlo and molecular dynamics simulations. It has been observed that in [n]collarenes the ratio of the interaction energies of a cation with it and the cation with the basic building unit (benzene) can be correlated to its ion selectivity. We find that collarenes are excellent and efficient ionophores that bind cations through cation-π interactions. [6]Collarene is found to be a selective host for Li+ and Mg2+, [8]collarene for K+ and Sr2+, and [10]collarene for Cs+ and Ba2+. This finding indicates that [10]collarene and [8]collarene could be used for effective separation of highly radioactive isotopes, 137Cs and 90Sr, which are major constituents of nuclear wastes. More interestingly, collarenes of larger cavity size can be useful in capturing organic cations. [12]Collarene exhibits a pronounced affinity for tetramethylammonium cation and acetylcholine, which implies that it could serve as a model for acetylcholinestrase. Thus, collarenes can prove to be novel and effective ionophores/model-receptors capable of heralding a new direction in molecular recognition and host-guest chemistry.

Molecular and functional characterization of a novel low-affinity cation transporter (LCT1) in higher plants

The transport of cations across membranes in higher plants plays an essential role in many physiological processes including mineral nutrition, cell expansion, and the transduction of environmental signals. In higher plants the coordinated expression of transport mechanisms is essential for specialized cellular processes and for adaptation to variable environmental conditions. To understand the molecular basis of cation transport in plant roots, a Triticum aestivum cDNA library was used to complement a yeast mutant deficient in potassium (K+) uptake. Two genes were cloned that complemented the mutant: HKT1 and a novel cDNA described in this report encoding a cation transporter, LCT1 (low-affinity cation transporter). Analysis of the secondary structure of LCT1 suggests that the protein contains 8–10 transmembrane helices and a hydrophilic amino terminus containing sequences enriched in Pro, Ser, Thr, and Glu (PEST). The transporter activity was assayed using radioactive isotopes in yeast cells expressing the cDNA. LCT1 mediated low-affinity uptake of the cations Rb+ and Na+, and possibly allowed Ca2+ but not Zn2+ uptake. LCT1 is expressed in low abundance in wheat roots and leaves. The precise functional role of this cation transporter is not known, although the competitive inhibition of cation uptake by Ca2+ has parallels to whole plant and molecular studies that have shown the important role of Ca2+ in reducing Na+ uptake and ameliorating Na+ toxicity. The structure of this higher plant ion transport protein is unique and contains PEST sequences.

Nitration of γ-tocopherol and oxidation of α-tocopherol by copper-zinc superoxide dismutase/H2O2/NO2−: Role of nitrogen dioxide free radical

Copper-zinc superoxide dismutase (Cu,ZnSOD) is the antioxidant enzyme that catalyzes the dismutation of superoxide (O2•−) to O2 and H2O2. In addition, Cu,ZnSOD also exhibits peroxidase activity in the presence of H2O2, leading to self-inactivation and formation of a potent enzyme-bound oxidant. We report in this study that lipid peroxidation of l-α-lecithin liposomes was enhanced greatly during the SOD/H2O2 reaction in the presence of nitrite anion (NO2−) with or without the metal ion chelator, diethylenetriaminepentacetic acid. The presence of NO2− also greatly enhanced α-tocopherol (α-TH) oxidation by SOD/H2O2 in saturated 1,2-dilauroyl-sn-glycero-3-phosphatidylcholine liposomes. The major product identified by HPLC and UV-studies was α-tocopheryl quinone. When 1,2-diauroyl-sn-glycero-3-phosphatidylcholine liposomes containing γ-tocopherol (γ-TH) were incubated with SOD/H2O2/NO2−, the major product identified was 5-NO2-γ-TH. Nitrone spin traps significantly inhibited the formation of α-tocopheryl quinone and 5-NO2-γ-TH. NO2− inhibited H2O2-dependent inactivation of SOD. A proposed mechanism of this protection involves the oxidation of NO2− by an SOD-bound oxidant to the nitrogen dioxide radical (•NO2). In this study, we have shown a new mechanism of nitration catalyzed by the peroxidase activity of SOD. We conclude that NO2− is a suitable probe for investigating the peroxidase activity of familial Amyotrophic Lateral Sclerosis-linked SOD mutants.

A single serine residue controls the cation dependence of substrate transport by the rat serotonin transporter

The serotonin transporter (SERT) is a member of the Na+/Cl−-dependent neurotransmitter transporter family and constitutes the target of several clinically important antidepressants. Here, replacement of serine-545 in the recombinant rat SERT by alanine was found to alter the cation dependence of serotonin uptake. Substrate transport was now driven as efficiently by LiCl as by NaCl without significant changes in serotonin affinity. Binding of the antidepressant [3H]imipramine occurred with 1/5th the affinity, whereas [3H]citalopram binding was unchanged. These results indicate that serine-545 is a crucial determinant of both the cation dependence of serotonin transport by SERT and the imipramine binding properties of SERT.

Proper sorting of the cation-dependent mannose 6-phosphate receptor in endosomes depends on a pair of aromatic amino acids in its cytoplasmic tail

The 67-amino acid cytoplasmic tail of the cation-dependent mannose 6-phosphate receptor (CD-MPR) contains a signal(s) that prevents the receptor from entering lysosomes where it would be degraded. To identify the key residues required for proper endosomal sorting, we analyzed the intracellular distribution of mutant forms of the receptor by Percoll density gradients. A receptor with a Trp19 → Ala substitution in the cytoplasmic tail was highly missorted to lysosomes whereas receptors with either Phe18 → Ala or Phe13 → Ala mutations were partially defective in avoiding transport to lysosomes. Analysis of double and triple mutants confirmed the key role of Trp19 for sorting of the CD-MPR in endosomes, with Phe18, Phe13, and several neighboring residues contributing to this function. The addition of the Phe18-Trp19 motif of the CD-MPR to the cytoplasmic tail of the lysosomal membrane protein Lamp1 was sufficient to partially impair its delivery to lysosomes. Replacing Phe18 and Trp19 with other aromatic amino acids did not impair endosomal sorting of the CD-MPR, indicating that two aromatic residues located at these positions are sufficient to prevent the receptor from trafficking to lysosomes. However, alterations in the spacing of the diaromatic amino acid sequence relative to the transmembrane domain resulted in receptor accumulation in lysosomes. These findings indicate that the endosomal sorting of the CD-MPR depends on the correct presentation of a diaromatic amino acid-containing motif in its cytoplasmic tail. Because a diaromatic amino acid sequence is also present in the cytoplasmic tail of other receptors known to be internalized from the plasma membrane, this feature may prove to be a general determinant for endosomal sorting.