scholarly journals Structural requirements for the utilization of ascorbate analogues in the prolyl 4-hydroxylase reaction

1994 ◽  
Vol 300 (1) ◽  
pp. 75-79 ◽  
Author(s):  
G Tschank ◽  
J Sanders ◽  
K H Baringhaus ◽  
F Dallacker ◽  
K I Kivirikko ◽  
...  
Keyword(s):  
Negative Charge ◽  
Substrate Inhibition ◽  
Side Chain ◽  
Structural Requirements ◽  
Response Behaviour ◽  
Structural Analogues ◽  
Microsomal Hydroxylation ◽  
Ring Position ◽  
Charged Group ◽  
Positively Charged

The ability of structural analogues of ascorbate to serve as substitutes for this reducing agent in the prolyl 4-hydroxylase reaction was studied. In experiments using the purified enzyme, variations of the compounds′ side chain were compatible with co-substrate activity. The presence of very large hydrophobic substituents or a positively charged group caused an increase in the observed Km values. A negative charge and smaller modifications did not change the affinity to the enzyme when compared with L-ascorbate. 6-Bromo-6-deoxy-L-ascorbate had a lower Km than the physiological reductant. Substitution at the -OH group in ring position 3 prevented binding to the enzyme. The same pattern of activity was observed when the full and uncoupled prolyl 4-hydroxylase reactions were studied. The Vmax. values with all compounds were similar. The reaction of microsomal prolyl 4-hydroxylase was supported by D-isoascorbate, O6-tosyl-L-ascorbate and 5-deoxy-L-ascorbate, giving the same dose-response behaviour as L-ascorbate itself. Again, 6-bromo-6-deoxy-L-ascorbate gave a lower Km and a similar Vmax. value. L-Ascorbic acid 6-carboxylate produced substrate inhibition at concentrations above 0.3 mM. The Km and Vmax. values calculated from concentrations up to 0.2 mM were similar to those of L-ascorbate. The enzyme activity observed with 6-amino-6-deoxy-L-ascorbate was very low in the microsomal hydroxylation system. The calculated Vmax. value was lower than that of L-ascorbate, suggesting a restriction of the access of this compound to the enzyme.

scholarly journals ANTIGEN RECOGNITION AND THE IMMUNE RESPONSE

1972 ◽  
Vol 136 (2) ◽  
pp. 387-391 ◽  
Author(s):  
Sefik S. Alkan ◽  
Maurice E. Bush ◽  
Danute E. Nitecki ◽  
Joel W. Goodman
Keyword(s):  
Immune Response ◽  
Hydroxyl Group ◽  
Side Chain ◽  
Low Molecular Weight ◽  
Carboxyl Group ◽  
Amino Group ◽  
Structural Requirements ◽  
Phenolic Ring ◽  
Cellular Immune ◽  
Charged Group

The low molecular weight compound L-tyrosine-azobenzenearsonate (RAT) induces a cellular immune response in guinea pigs. The contribution of the side chain of tyrosine to the immunogenicity of RAT and the structural requirements at that position for immunogenicity were assessed by synthesizing a series of analogs of RAT containing modifications in the side chain of tyrosine and employing them as immunogens. Removal of either the carboxyl or amino group did not markedly affect immunogenicity, measured by the induction of delayed cutaneous sensitivity, whereas deletion of both completely abolished it. However, a charged group was not required since side chains containing a polar hydroxyl group could substitute for chains bearing an amino or carboxyl group. The size of the side chain exerted a pronounced influence; the charged or polar substituent had to be extended from the phenolic ring by at least two carbon atoms in order to confer immunogenicity.

Enhanced the Negative Charges of Antheraea pernyi Silk Fibroin by Methylglyoxal Modification

2015 ◽  
Vol 1101 ◽  
pp. 307-313 ◽  
Author(s):  
Jing Li ◽  
Ceng Zhang ◽  
Yi Zhang ◽  
Yan Ni Yu ◽  
Jing Wan Luo ◽  
...  
Keyword(s):  
Aqueous Solution ◽  
Silk Fibroin ◽  
Nmr Spectra ◽  
Negative Charge ◽  
Controlled Drug Release ◽  
Side Chain ◽  
Antheraea Pernyi ◽  
H Nmr ◽  
Chemically Modified ◽  
Positively Charged

Antheraea pernyi silk fibroin has favorable biocompatibility, good bioactivity and controllable biodegradability, meeting the basic requirements of controlled drug release carriers. Enhancing the negative charge of silk fibroin could further increase the encapsulation and loading efficiency of positively charged drugs. In this study, Antheraea pernyi silk fibroin was chemically modified by methylglyoxal in aqueous solution. The electric charge properties of Antheraea pernyi silk fibroin were examined to characterize the modification, the results indicated that the isoelectric point of Antheraea pernyi silk fibroin decreased from 4.5 to 3.9, and the zeta potential reduced from-11.7 mV to-12.8 mV. Amino acid analysis and 1H-NMR spectra showed that arginine residue of Antheraea pernyi silk fibroin side chain was modified by methylglyoxal for enhancing negative charge of silk fibroin. These results suggested that methylglyoxal-modified Antheraea pernyi silk fibroin could be considered as a potential starting material in loading positively charged drugs.

scholarly journals Lysine carbamylation for enzymatic function: metal and structural requirements

2014 ◽  
Vol 70 (a1) ◽  
pp. C301-C301
Author(s):  
Yin-Cheng Hsieh ◽  
Sunney Chan ◽  
Yuh-Shyong Yang ◽  
Chun-Jung Chen
Keyword(s):  
Negative Charge ◽  
Iminodiacetic Acid ◽  
Enzymatic Activities ◽  
Metal Coordination ◽  
Side Chain ◽  
Post Translational Modification ◽  
Structural Requirements ◽  
Enzymatic Function ◽  
Tetraodon Nigroviridis

Lysine carbamylation, a post-translational modification, facilitates metal coordination for specific enzymatic activities. Carbamylation on lysine extends the residue length by ~2 Å and changes the side chain from a positive to negative charge at neutral pH. The proteins involved with lysine carbamylation are found to be related to the human diseases, such as type 2 diabetes, developmental delay, metabolic acidosis, mental retardation, hypotonia and seizures. We have determined structures of the vertebrate dihydropyrimidinase from Tetraodon nigroviridis (TnDhp) in various states: the apo enzyme as well as two forms of the holo enzyme with one and two metals at the catalytic site. The essential active-site structural requirements have been identified with possible existence of four metal-mediated stages of lysine carbamylation. Only one metal is sufficient for stabilizing lysine carbamylation; however, the post-translational lysine carbamylation facilitates additional metal coordination for the regulation of specific enzymatic activities through controlling the conformations of two dynamic loops, Ala69–Arg74 and Met158–Met165, located in the tunnel for the substrate entrance. The substrate/product tunnel is in the "open form" in the apo-TnDhp, in the "intermediate state" in the mono-metal TnDhp, and in the "closed form" in the di-metal TnDhp structure, respectively. Structural comparison also suggests that the C-terminal tail plays a role in the enzymatic function through interactions with the Ala69–Arg74 dynamic loop. In addition, the structures of the di-metal TnDhp in complexes with hydantoin, N-carbamyl-β-alanine and N-carbamyl-β-amino isobutyrate, as well as apo-TnDhp in complex with a product analog, N-(2-acetamido)-iminodiacetic acid, have been determined. These structural results illustrate how a protein exploits unique lysines and the metal distribution to accomplish lysine carbamylation as well as subsequent enzymatic functions.

Replacement of the interchain disulfide bridge-forming amino acids A7 and B7 by glutamate impairs the structure and activity of insulin

2004 ◽  
Vol 385 (12) ◽  
pp. 1171-1175 ◽  
Author(s):  
Zhan-Yun Guo ◽  
Xiao-Yuan Jia ◽  
You-Min Feng
Keyword(s):  
Biological Activity ◽  
Disulfide Bonds ◽  
Negative Charge ◽  
Disulfide Bridge ◽  
Site Directed Mutagenesis ◽  
Side Chain ◽  
Insulin Analogs ◽  
High Biological Activity ◽  
Chemical Groups ◽  
Structure And Activity

Abstract Insulin contains three disulfide bonds, one intrachain bond, A6–A11, and two interchain bonds, A7–B7 and A20–B19. Site-directed mutagenesis results (the two cysteine residues of disulfide A7–B7 were replaced by serine) showed that disulfide A7–B7 is crucial to both the structure and activity of insulin. However, chemical modification results showed that the insulin analogs still retained relatively high biological activity when A7Cys and B7Cys were modified by chemical groups with a negative charge. Did the negative charge of the modification groups restore the loss of activity and/or the disturbance of structure of these insulin analogs caused by deletion of disulfide A7–B7? To answer this question, an insulin analog with both A7Cys and B7Cys replaced by Glu, which has a long side-chain and a negative charge, was prepared by protein engineering, and its structure and activity were analyzed. Both the structure and activity of the present analog are very similar to that of the mutant with disulfide A7–B7 replaced by Ser, but significantly different from that of wild-type insulin. The present results suggest that removal of disulfide A7–B7 will result in serious loss of biological activity and the native conformation of insulin, even if the disulfide is replaced by residues with a negative charge.

First Principles Investigation of the Stability and the Chemical Behavior of Hydrogen in ThCoH4

2011 ◽  
Vol 66 (3) ◽  
pp. 269-274
Author(s):  
Samir F. Matar
Keyword(s):  
Negative Charge ◽  
Geometry Optimization ◽  
Full Geometry Optimization ◽  
Intermetallic Matrix ◽  
Charge Analysis ◽  
The Stability ◽  
The One ◽  
Site Selective ◽  
Bader Charge Analysis ◽  
Positively Charged

We address the changes in the electronic structure brought by the insertion of hydrogen into ThCo leading to the experimentally observed ThCoH4. Full geometry optimization positions the hydrogen in three sites stabilized in the expanded intermetallic matrix. From a Bader charge analysis, hydrogen is found to be in a narrow iono-covalent (~−0.6) to covalent (~−0.3) bonding which should enable site-selective desorption. The overall chemical picture shows a positively charged Thδ+ with the negative charge redistributed over a complex anion {CoH4}δ− with δ~1.8. Nevertheless this charge transfer remains far from the one in the more ionic hydridocobaltate anion CoH54− in Mg2CoH5, due to the largely electropositive character of Mg.

Mutation of a single amino acid converts the human water channel aquaporin 5 into an anion channel

2013 ◽  
Vol 305 (6) ◽  
pp. C663-C672 ◽  
Author(s):  
Xue Qin ◽  
Walter F. Boron
Keyword(s):  
Water Channel ◽  
Anion Channel ◽  
Single Amino Acid ◽  
Side Chain ◽  
Wild Type ◽  
Aquaporin 5 ◽  
Anion Conductance ◽  
Electrode Voltage ◽  
Microelectrode Measurements ◽  
Positively Charged

Aquaporin 6 (AQP6) is unique among mammalian AQPs in being an anion channel with negligible water permeability. However, the point mutation Asn60Gly converts AQP6 from an anion channel into a water channel. In the present study of human AQP5, we mutated Leu51 (corresponding to residue 61 in AQP6), the side chain of which faces the central pore. We evaluated function in Xenopus oocytes by two-electrode voltage clamp, video measurements of osmotic H2O permeability ( Pf), microelectrode measurements of surface pH (pHS) to assess CO2 permeability, and surface biotinylation. We found that AQP5-L51R does not exhibit the H2O or CO2 permeability of the wild-type protein but instead has a novel p-chloromercuribenzene sulfonate (pCMBS)-sensitive current. The double mutant AQP5-L51R/C182S renders the conductance insensitive to pCMBS, demonstrating that the current is intrinsic to AQP5. AQP5-L51R has the anion permeability sequence I− > NO3− ≅ NO2− > Br− > Cl− > HCO3− > gluconate. Of the other L51 mutants, L51T (polar uncharged) and L51V (nonpolar) retain H2O and CO2 permeability and do not exhibit anion conductance. L51D and L51E (negatively charged) have no H2O or CO2 permeability. L51K (positively charged) has an intermediate H2O and CO2 permeability and anion conductance. L51H is unusual in having a relatively low CO2 permeability and anion conductance, but a moderate Pf. Thus, positively charged mutations of L51 can convert AQP5 from a H2O/CO2 channel into an anion channel. However, the paradoxical effect of L51H is consistent with the hypothesis that CO2, in part, takes a pathway different from H2O through AQP5.

scholarly journals Structure of a Talaromyces pinophilus GH62 arabinofuranosidase in complex with AraDNJ at 1.25 Å resolution

2018 ◽  
Vol 74 (8) ◽  
pp. 490-495 ◽  
Author(s):  
Olga V. Moroz ◽  
Lukasz F. Sobala ◽  
Elena Blagova ◽  
Travis Coyle ◽  
Wei Peng ◽  
...  
Keyword(s):  
Plant Biomass ◽  
Cellulosic Biomass ◽  
Structural Basis ◽  
Side Chain ◽  
Polymeric Substrates ◽  
Hydrolysis Of ◽  
Insight Into ◽  
Positively Charged ◽  
Talaromyces Pinophilus ◽  
Resolution Structure

The enzymatic hydrolysis of complex plant biomass is a major societal goal of the 21st century in order to deliver renewable energy from nonpetroleum and nonfood sources. One of the major problems in many industrial processes, including the production of second-generation biofuels from lignocellulose, is the presence of `hemicelluloses' such as xylans which block access to the cellulosic biomass. Xylans, with a polymeric β-1,4-xylose backbone, are frequently decorated with acetyl, glucuronyl and arabinofuranosyl `side-chain' substituents, all of which need to be removed for complete degradation of the xylan. As such, there is interest in side-chain-cleaving enzymes and their action on polymeric substrates. Here, the 1.25 Å resolution structure of the Talaromyces pinophilus arabinofuranosidase in complex with the inhibitor AraDNJ, which binds with a K d of 24 ± 0.4 µM, is reported. Positively charged iminosugars are generally considered to be potent inhibitors of retaining glycosidases by virtue of their ability to interact with both acid/base and nucleophilic carboxylates. Here, AraDNJ shows good inhibition of an inverting enzyme, allowing further insight into the structural basis for arabinoxylan recognition and degradation.

THE EFFECT OF PROCOAGULANT PHOSPHOLIPID VESICLES WITH NET POSITIVE CHARGE ON THE ACTIVITY OF PROTHROMBINASE

1987 ◽  
Author(s):  
J Rosing ◽  
H Speijer ◽  
J W P Govers-Riemslag ◽  
R F A Zwaal
Keyword(s):  
Vitamin K ◽  
Negative Charge ◽  
Positive Charge ◽  
Coagulation Factor ◽  
Phospholipid Vesicles ◽  
Electrostatic Model ◽  
Acidic Phospholipid ◽  
Positively Charged ◽  
Net Positive Charge ◽  
Prothrombin Activation

It is generally thought that procoagulant phospholipid surfaces that promote the activation of vitamin K-dependent coagulation factors should have a net negative charge in order to promote calcium-dependent binding of the enzymes (FVIIa, FIXa and FXa) and substrates (prothrombin and FX) of the coagulation factor-activating complexes. Two models have been proposed to explain calcium-mediated association of vitamin K-dependent proteins with phospholipid: a) an electrostatic model, in which a positively-charged protein-calcium complex is attracted by a negatively-charged phospholipid surface and b) a chelation model in which a coordination complex is formed between calcium ions, γ-carboxyglutamic acids of the proteins and negatively-charged membrane phospholipids. To study the effect of the electrostatic potential of phospholipid vesicles on their activity in the pro-thrombinase complex the net charge of vesicles was varied by introduction of varying amounts of positively-charged stearylamine in the membrane surface. Introduction of 0-15 mole% stearylamine in phospholipid vesicles that contained 5 mole% phosphatidylseri-ne (PS) hardly affected their activity in prothrombin activation. Electrophoretic analysis showed that vesicles with > 5 mole% stearylamine had a net positive charge. The procoagulant activity of vesicles that contained phosphatidic acid, phosphatidylglyce-rol, phosphatidylinositol or phosphatidyl-glactate (PLac) as acidic phospholipid was much more effected by incorporation of stearylamine. Amounts of stearylamine that compensated the negative charge of acidic phospholipid caused considerable inhibition of the activity of the latter vesicles in prothrombin activation. The comparison of vesicles containing PS and PLac as acidic phospholipid is of special interest. PS and PLac only differ by the presence of NH+ 3-group in the serine moiety of PS. Thus, in spite of the fact that vesicles with PLac are more negatively charged than vesicles with PS, they are less procoagulant. Our results show that a) although procoagulant membranes have to contain acidic phospholipids there is no requirement for a net negative charge, b) the amino group of phosphatidylserine has an important function in the interaction of procoagulant membranes with vitamin K-dependent proteins and c) the chelation model can satisfactorily explain calcium-mediated lipid-protein association.

scholarly journals The Position of Solid Carbon Dioxide in the Triboelectric Series

2019 ◽  
Vol 72 (8) ◽  
pp. 633 ◽  
Author(s):  
Jinyang Zhang ◽  
Simone Ciampi
Keyword(s):  
Carbon Dioxide ◽  
Negative Charge ◽  
Plastic Material ◽  
Indirect Evidence ◽  
Liquid Carbon ◽  
Solid Carbon Dioxide ◽  
Dry Ice ◽  
First Time ◽  
Charge Sign ◽  
Positively Charged

The process of releasing liquid carbon dioxide from a fire extinguisher is accompanied by a strong static charging of the plastic material making up the extinguisher discharge horn. Firefighters often report an electric shock when operating CO2 extinguishers, but the origin of this electrostatic hazard is largely unknown. Here, we begin to investigate this phenomenon, and test the hypothesis of plastic samples being tribocharged on contact with rapidly flowing solid CO2. Using Faraday pail measurements, we show that non-conductive polymers gain a net static charge when brought in and out of contact with dry ice (solid CO2). These measurements of charge sign and magnitude give indirect evidence helping to place solid CO2 for the first time on the triboelectric series. Polydimethylsiloxane (PDMS), polytetrafluoroethylene (PTFE), and polyvinyl chloride (PVC) samples acquire a negative charge when rubbed against dry ice, whereas poly(methyl methacrylate) (PMMA), glass, and nylon surfaces become positively charged. Therefore, we suggest the position of dry ice in the triboelectric series to be close to that of materials with stable cations and unstable anions, possibly locating it between PMMA and PVC.

A study of the relationships of interactions between Asp-201, Na+ or K+, and galactosyl C6 hydroxyl and their effects on binding and reactivity of β-galactosidase

2004 ◽  
Vol 82 (2) ◽  
pp. 275-284 ◽  
Author(s):  
Julia Xu ◽  
Mary A.A McRae ◽  
Scott Harron ◽  
Beatrice Rob ◽  
Reuben E Huber
Keyword(s):  
Physical Properties ◽  
Negative Charge ◽  
Binding Pocket ◽  
Side Chain ◽  
Wild Type ◽  
Sodium Potassium ◽  
The Difference ◽  
Potassium Binding ◽  
Covalent Intermediate ◽  
Catalytic Effects

The interactions between Na+ (and K+) and Asp-201 of β-galactosidase were studied. Analysis of the changes in Km and Vmax showed that the Kd for Na+ of wild type β-galactosidase (0.36 ± 0.09 mM) was about 10× lower than for K+ (3.9 ± 0.6 mM). The difference is probably because of the size and other physical properties of the ions and the binding pocket. Decreases of Km as functions of Na+ and K+ for oNPG and pNPG and decreases of the Ki of both shallow and deep mode inhibitors were similar, whereas the Km and Ki of substrates and inhibitors without C6 hydroxyls remained constant. Thus, Na+ and K+ are important for binding galactosyl moieties via the C6 hydroxyl throughout catalysis. Na+ and K+ had lesser effects on the Vmax. The Vmax of pNPF and pNPA (substrates that lack a C6 hydroxyl) did not change upon addition of Na+ or K+, showing that the catalytic effects are also mediated via the C6 hydroxyl. Arrhenius plots indicated that Na+, but not K+, caused k3 (degalactosylation) to increase. Na+ also caused the k2 (galactosylation) with oNPG, but not with pNPG, to increase. In contrast, K+ caused the k2 values with both oNPG and pNPG to increase. Na+ and K+ mainly altered the entropies of activation of k2 and k3 with only small effects on the enthalpies of activation. This strongly suggests that only the positioning of the substrate, transition states, and covalent intermediate are altered by Na+ and K+. Further evidence that positioning is important was that substitution of Asp-201 with a Glu caused the Km and Ki values to increase significantly. In addition, the Kd values for Na+ or K+ were 5 to 8 fold higher. The negative charge of Asp-201 was shown to be vital for Na+ and K+ binding. Large amounts of Na+ or K+ had no effect on the very large Km and Ki values of D201N-β-galactosidase and the Vmax values changed minimally and in a linear rather than hyperbolic way. D201F-β-galactosidase, with a very bulky hydrophobic side chain in place of Asp, essentially obliterated all binding and catalysis.Key words: β-galactosidase, sodium, potassium, binding, aspartic acid.

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