scholarly journals PtdIns4P-mediated electrostatic forces influence S-acylation of peripheral proteins at the Golgi complex

2020 ◽  
Vol 40 (1) ◽  
Author(s):  
Sabrina Chumpen Ramirez ◽  
Micaela R. Astrada ◽  
Jose L. Daniotti
Keyword(s):  
Amino Acids ◽  
Golgi Complex ◽  
Electrostatic Interactions ◽  
Transmembrane Protein ◽  
Basic Amino Acid ◽  
Soluble Proteins ◽  
Post Translational Modification ◽  
Hydrophobic Region ◽  
Membrane Interaction ◽  
Basic Amino Acids

Abstract Protein S-acylation is a reversible post-translational modification involving the addition of fatty acids to cysteines and is catalyzed by transmembrane protein acyltransferases (PATs) mainly expressed at the Golgi complex. In case of soluble proteins, S-acylation confers stable membrane attachment. Myristoylation or farnesylation of many soluble proteins constitutes the initial transient membrane adsorption step prior to S-acylation. However, some S-acylated soluble proteins, such as the neuronal growth-associated protein Growth-associated protein-43 (GAP-43), lack the hydrophobic modifications required for this initial membrane interaction. The signals for GAP-43 S-acylation are confined to the first 13 amino acids, including the S-acylatable cysteines 3 and 4 embedded in a hydrophobic region, followed by a cluster of basic amino acids. We found that mutation of critical basic amino acids drastically reduced membrane interaction and hence S-acylation of GAP-43. Interestingly, acute depletion of phosphatidylinositol 4-phosphate (PtdIns4P) at the Golgi complex reduced GAP-43 membrane binding, highlighting a new, pivotal role for this anionic lipid and supporting the idea that basic amino acid residues are involved in the electrostatic interactions between GAP-43 and membranes of the Golgi complex where they are S-acylated.

Basic amino acid induced isomerization of a spiropyran: towards visual recognition of basic amino acids in water

2007 ◽  
Vol 31 (11) ◽  
pp. 1878 ◽  
Author(s):  
Yuanyuan Liu ◽  
Meigong Fan ◽  
Shuxiao Zhang ◽  
Xiaohai Sheng ◽  
Jiannian Yao
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Visual Recognition ◽  
Basic Amino Acid ◽  
Basic Amino Acids

Influence of organic cations on basic amino-acid uptake by human placental villi

1995 ◽  
Vol 7 (6) ◽  
pp. 1491
Author(s):  
RB Krishna ◽  
J Dancis ◽  
M Levitz
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Basic Amino Acid ◽  
Amino Acid Uptake ◽  
Progressive Increase ◽  
Amino Acid Transporters ◽  
Acid Uptake ◽  
Chorionic Villi ◽  
Basic Amino Acids ◽  
Kinetics Of

Human placental chorionic villi were incubated for 30 min with [3H]lysine or [3H]arginine and the distribution ratios (intracellular:extracellular concentrations) were determined. The ratios remained unchanged when Na+ in Earle's buffered salt solution was replaced with Li+. When Na+ was replaced with choline there was a significant increase is distribution ratios (lysine 1.34 +/- 0.33 v. 3.99 +/- 0.15, arginine 1.95 +/- 0.37 v. 5.05 +/- 1.16). Leucine, a neutral amino acid with a Na(+)-independent transport system, was unaffected by choline substitution. The distribution ratio for alanine, which is Na(+)-dependent, was reduced (2.50 +/- 0.41 v. 1.45 +/- 0.20). Two other quarternary amines, acetyl-beta-methylcholine and tetraethylammonium chloride (TEA) caused similar increases in the distribution ratios of the basic amino acids. Hordenine, a tertiary amine, was less effective and there was little or no effect with ephedrine, a secondary amine. The choline effect was first observable at concentrations of 105 mM. With TEA, there was a progressive increase in distribution ratios beginning at 29 mM. Lysine efflux was measured after incubation of villi with lysine in Earle's buffer or choline buffer. Lysine was rapidly released to the fresh medium with 25% more retained in choline-exposed villi. The amines may cause alterations in the kinetics of basic amino-acid transporters or may modify other aspects of placental physiology permitting an increase retention of the basic amino acids.

scholarly journals The Acinetobacter Outer Membrane Contains Multiple Specific Channels for Carbapenem β-Lactams as Revealed by Kinetic Characterization Analyses of Imipenem Permeation into Acinetobacter baylyi Cells

2017 ◽  
Vol 61 (3) ◽  
Author(s):  
Jorgelina Morán-Barrio ◽  
María M. Cameranesi ◽  
Verónica Relling ◽  
Adriana S. Limansky ◽  
Luciano Brambilla ◽  
...  
Keyword(s):  
Amino Acids ◽  
Outer Membrane ◽  
Binding Sites ◽  
Carbon Sources ◽  
Basic Amino Acid ◽  
Acinetobacter Baylyi ◽  
Content Type ◽  
Whole Cells ◽  
Basic Amino Acids ◽  
Uptake Sites

ABSTRACT The number and type of outer membrane (OM) channels responsible for carbapenem uptake in Acinetobacter are still not well defined. Here, we addressed these questions by using Acinetobacter baylyi as a model species and a combination of methodologies aimed to characterize OM channels in their original membrane environment. Kinetic and competition analyses of imipenem (IPM) uptake by A. baylyi whole cells allowed us to identify different carbapenem-specific OM uptake sites. Comparative analyses of IPM uptake by A. baylyi wild-type (WT) cells and ΔcarO mutants lacking CarO indicated that this OM protein provided a carbapenem uptake site displaying saturable kinetics and common binding sites for basic amino acids compatible with a specific channel. The kinetic analysis uncovered another carbapenem-specific channel displaying a somewhat lower affinity for IPM than that of CarO and, in addition, common binding sites for basic amino acids as determined by competition studies. The use of A. baylyi gene deletion mutants lacking OM proteins proposed to function in carbapenem uptake in Acinetobacter baumannii indicated that CarO and OprD/OccAB1 mutants displayed low but consistent reductions in susceptibility to different carbapenems, including IPM, meropenem, and ertapenem. These two mutants also showed impaired growth on l-Arg but not on other carbon sources, further supporting a role of CarO and OprD/OccAB1 in basic amino acid and carbapenem uptake. A multiple-carbapenem-channel scenario may provide clues to our understanding of the contribution of OM channel loss or mutation to the carbapenem-resistant phenotype evolved by pathogenic members of the Acinetobacter genus.

scholarly journals Crystallization of recombinant hemoglobins with basic amino acid substitutions (Lys and Arg) at the beta 6 position

Blood ◽  
1994 ◽  
Vol 84 (4) ◽  
pp. 1309-1313
Author(s):  
K Adachi ◽  
CH Lai ◽  
P Konitzer ◽  
M Donahee ◽  
A Campbell ◽  
...  
Keyword(s):  
Amino Acids ◽  
Expression System ◽  
Basic Amino Acid ◽  
Cellulose Acetate Electrophoresis ◽  
Yeast Expression System ◽  
Basic Amino Acids ◽  
Beta 2 ◽  
Hb Concentration ◽  
Hb C

We have produced recombinant hemoglobins (rHbs) alpha 2 beta 2(6Glu-- >Lys) (rHb beta E6K) and alpha 2 beta 2(6Glu-->Arg) (rHb beta E6R) using a yeast expression system coupled with a polymerase chain reaction (PCR)-based mutagenesis strategy for studies focused on defining determinants that facilitate crystallization of Hb C (alpha 2 beta 2(6Lys)). rHb beta E6K had the same electrophoretic mobility as native human Hb C, whereas rHb beta E6R migrated slightly slower than Hb C on cellulose acetate electrophoresis. The carbonmonoxy (CO) forms of rHb beta E6K and rHb beta E6R formed tetrahedral crystals in vitro in 2.3 mol/L phosphate buffer just like native Hb C. The Hb concentration required for crystallization of CO-rHb beta E6R was lower than that of CO-rHb beta E6K, suggesting that stronger basic amino acids at the beta 6 position accelerate crystallization of Hb. However, the size of rHb beta E6R crystals was smaller than that of rHb beta E6K. Crystallization of native Hb C and both rHbs was inhibited by Hb F. These results suggest that alpha 2 beta gamma-heterohybrids that have basic amino acids at the beta 6 position behave similarly and are unable to crystallize like Hb C.

Basic amino acid transport in renal papilla: microinfusion of Henle's loops and vasa recta

1993 ◽  
Vol 265 (6) ◽  
pp. F830-F838 ◽  
Author(s):  
W. H. Dantzler ◽  
S. Silbernagl
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Transport ◽  
Basic Amino Acid ◽  
Renal Papilla ◽  
Tubular Structures ◽  
Acid Transport ◽  
Basic Amino Acids ◽  
Neutral Amino Acids ◽  
Vasa Recta

To determine whether basic amino acids, like acidic and neutral amino acids, could be reabsorbed distal to tips of Henle's loops and recycled between loops and vasa recta in the renal papilla, we continuously microinfused ascending Henle's loops and vasa recta with 14C-labeled L-lysine (L-Lys; 1.28 mM) or L-arginine (L-Arg; 1.17 mM) and 3H-labeled inulin. We also determined percent of recovered radiolabel as intact amino acid. Like acidic and neutral amino acids, relative to inulin, approximately 30% of L-Lys and approximately 45% of L-Arg microinfused into Henle's loops were reabsorbed. However, whereas radiolabeled L-Lys reabsorption, like reabsorption of acidic and neutral amino acids, was not readily inhibited, radiolabeled L-Arg reabsorption was reduced to approximately 25% by addition of unlabled L-Arg (50 mM) or L-homoarginine (L-Homo-Arg) (50 mM) to infusate. This observation provides greater evidence for specific, carrier-mediated reabsorption for L-Arg than for acidic or neutral amino acids. About 36% (relative to inulin) of each of these amino acids microinfused into ascending vasa recta apparently was transferred directly into ipsilateral tubular structures (probably thin descending limbs of Henle's loops). Transfer of radiolabeled L-Arg was reduced to approximately 8% by the inclusion of unlabeled L-Arg (50 mM) in infusate. Transfer of unlabeled L-Lys was unaffected by inclusion of unlabeled L-Lys (50 mM) in infusate but was reduced to approximately 20% by inclusion of unlabeled L-Arg (50 mM) or L-Homo-Arg (50 mM) in infusate. (ABSTRACT TRUNCATED AT 250 WORDS)

High sensitivity of Xenopus gustatory receptors to amino acids and bitter substances

1982 ◽  
Vol 243 (1) ◽  
pp. R42-R48 ◽  
Author(s):  
K. Yoshii ◽  
C. Yoshii ◽  
Y. Kobatake ◽  
K. Kurihara
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Xenopus Laevis ◽  
High Sensitivity ◽  
Basic Amino Acid ◽  
Gustatory Receptors ◽  
Basic Amino Acids ◽  
Neutral Amino Acids ◽  
Gustatory Nerve ◽  
Cross Adaptation

The gustatory nerve responses of the aquatic toad Xenopus laevis to salts, acids, amino acids, and bitter substances were recorded. 1) The gustatory receptors were sensitive to amino acids and bitter substances. The thresholds were 10(-7) M for Arg, 3 X 10(-9) M for strychnine, and 3 X 10(-8) M for quinine, 200-20,000 times lower than the thresholds for the respective stimuli in the bullfrog. 2) The basic and the neutral amino acids were effective whereas the acidic ones were not. Relations between the responses and log stimulus concentrations for the basic amino acids were linear in a wide concentration range whereas those for the neutral ones were of S shape. Cross-adaptation did not occur among pairs of a basic amino acid and a neutral one. 3) Responses to the basic amino acids and the basic bitter substances were suppressed by the addition of salts to the stimulating solutions, while those to the neutral and the acidic substances were not suppressed.

scholarly journals Mouse Ten-m/Odz Is a New Family of Dimeric Type II Transmembrane Proteins Expressed in Many Tissues

1999 ◽  
Vol 145 (3) ◽  
pp. 563-577 ◽  
Author(s):  
Toshitaka Oohashi ◽  
Xiao-Hong Zhou ◽  
Kang Feng ◽  
Brigitta Richter ◽  
Matthias Mörgelin ◽  
...  
Keyword(s):  
Amino Acids ◽  
Alkaline Phosphatase ◽  
Transmembrane Protein ◽  
Transmembrane Proteins ◽  
Extracellular Domain ◽  
Electrophoretic Analysis ◽  
Type Ii ◽  
Mrna Isoforms ◽  
Hydrophobic Region ◽  
Hydrophobic Domain

The Drosophila gene ten-m/odz is the only pair rule gene identified to date which is not a transcription factor. In an attempt to analyze the structure and the function of ten-m/odz in mouse, we isolated four murine ten-m cDNAs which code for proteins of 2,700–2,800 amino acids. All four proteins (Ten-m1–4) lack signal peptides at the NH2 terminus, but contain a short hydrophobic domain characteristic of transmembrane proteins, 300–400 amino acids after the NH2 terminus. About 200 amino acids COOH-terminal to this hydrophobic region are eight consecutive EGF-like domains. Cell transfection, biochemical, and electronmicroscopic studies suggest that Ten-m1 is a dimeric type II transmembrane protein. Expression of fusion proteins composed of the NH2-terminal and hydrophobic domain of ten-m1 attached to the alkaline phosphatase reporter gene resulted in membrane-associated staining of the alkaline phosphatase. Electronmicroscopic and electrophoretic analysis of a secreted form of the extracellular domain of Ten-m1 showed that Ten-m1 is a disulfide-linked dimer and that the dimerization is mediated by EGF-like modules 2 and 5 which contain an odd number of cysteines. Northern blot and immunohistochemical analyses revealed widespread expression of mouse ten-m genes, with most prominent expression in brain. All four ten-m genes can be expressed in variously spliced mRNA isoforms. The extracellular domain of Ten-m1 fused to an alkaline phosphatase reporter bound to specific regions in many tissues which were partially overlapping with the Ten-m1 immunostaining. Far Western assays and electronmicroscopy demonstrated that Ten-m1 can bind to itself.

scholarly journals Identification and Characterization of Heparin Binding Regions of the Fim2 Subunit of Bordetella pertussis

1998 ◽  
Vol 66 (5) ◽  
pp. 2256-2263 ◽  
Author(s):  
Cecile A. W. Geuijen ◽  
Rob J. L. Willems ◽  
Peter Hoogerhout ◽  
Wouter C. Puijk ◽  
Rob H. Meloen ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bordetella Pertussis ◽  
Sequence Similarity ◽  
Basic Amino Acid ◽  
Cross Reactivity ◽  
Site Directed Mutagenesis ◽  
Heparin Binding ◽  
Basic Amino Acids ◽  
Binding Regions

ABSTRACT Bordetella pertussis fimbriae bind to sulfated sugars such as heparin through the major subunit Fim2. The Fim2 subunit contains two regions, designated H1 and H2, which show sequence similarity with heparin binding regions of fibronectin, and the role of these regions in heparin binding was investigated with maltose binding protein (MBP)-Fim2 fusion proteins. Deletion derivatives of MBP-Fim2 showed that both regions are important for binding to heparin. The role of H2 in heparin binding was confirmed by site-directed mutagenesis in which basic amino acids were replaced by alanine. These studies revealed that Lys-186 and Lys-187 are important for heparin binding of MBP-Fim2, whereas Arg-179 is not required. Peptides derived from H1 and H2 (pepH1 and pepH2) also showed heparin binding activity. Using a series of peptides, in each of which a different basic amino acid was substituted for alanine, we demonstrated that the structural requirements for heparin binding differ significantly among pepH1 and pepH2 peptides. A Pepscan analysis of Fim2 revealed regions outside H1 and H2 which bind heparin and showed that not only basic amino acids but also tyrosines may be important for binding to sulfated sugars. A comparison of the heparin binding regions of Fim2 with homologous regions of Fim3 and FimX, two closely related but antigenically distinct fimbrial subunits, showed that basic amino acids and tyrosines are generally conserved. The major heparin binding regions identified in Fim2 are part of epitopes recognized by human antibodies, suggesting that the heparin binding regions are exposed at the fimbrial surface and are immunodominant. Since B. pertussis fimbriae show weak serological cross-reactivity, the differences in primary structure in the heparin binding regions of Fim2, Fim3, and FimX may affect antibody binding but not heparin binding, allowing the bacteria to evade antibody-mediated immunity by switching the fimbrial gene expressed.

scholarly journals Crystallization of recombinant hemoglobins with basic amino acid substitutions (Lys and Arg) at the beta 6 position

Blood ◽  
1994 ◽  
Vol 84 (4) ◽  
pp. 1309-1313 ◽  
Author(s):  
K Adachi ◽  
CH Lai ◽  
P Konitzer ◽  
M Donahee ◽  
A Campbell ◽  
...  
Keyword(s):  
Amino Acids ◽  
Expression System ◽  
Basic Amino Acid ◽  
Cellulose Acetate Electrophoresis ◽  
Yeast Expression System ◽  
Basic Amino Acids ◽  
Beta 2 ◽  
Hb Concentration ◽  
Hb C

Abstract We have produced recombinant hemoglobins (rHbs) alpha 2 beta 2(6Glu-- >Lys) (rHb beta E6K) and alpha 2 beta 2(6Glu-->Arg) (rHb beta E6R) using a yeast expression system coupled with a polymerase chain reaction (PCR)-based mutagenesis strategy for studies focused on defining determinants that facilitate crystallization of Hb C (alpha 2 beta 2(6Lys)). rHb beta E6K had the same electrophoretic mobility as native human Hb C, whereas rHb beta E6R migrated slightly slower than Hb C on cellulose acetate electrophoresis. The carbonmonoxy (CO) forms of rHb beta E6K and rHb beta E6R formed tetrahedral crystals in vitro in 2.3 mol/L phosphate buffer just like native Hb C. The Hb concentration required for crystallization of CO-rHb beta E6R was lower than that of CO-rHb beta E6K, suggesting that stronger basic amino acids at the beta 6 position accelerate crystallization of Hb. However, the size of rHb beta E6R crystals was smaller than that of rHb beta E6K. Crystallization of native Hb C and both rHbs was inhibited by Hb F. These results suggest that alpha 2 beta gamma-heterohybrids that have basic amino acids at the beta 6 position behave similarly and are unable to crystallize like Hb C.

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