Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Modifier”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,675 records · Page 93Linked to original sources
PubMed1975

Identification of the lysine residue modified during the activation of acetimidylation of horse liver alcohol dehydrogenase.

A single amino group in horse liver alcohol dehydrogenase was modified with methyl(14C)acetimidate by a differential labeling procedure. Lysine residues outside the active site were modified with ethyl acetimidate while a lysine residue in the active site was protected by the formation of an enzyme-NAD+-pyrazole complex. After the protecting reagents were removed, the enzyme was treated with methyl(14C)acetimidate. Enzyme activity was enhanced 13-fold as 1.1 (14C)acetimidyl group was incorporated per active site. A labeled peptide was isolated from a tryptic-chymotryptic digest of the modified enzyme in 35% overall yield. Amino acid composition and sequential Edman degradations identified the peptide as residues 219-229; lysine residue 228 was modified with the radioactive acetimidyl group.

Acetates↗
PubMed2005

Lantibiotic structures as guidelines for the design of peptides that can be modified by lantibiotic enzymes.

Lantibiotics are (methyl)lanthionine-containing bacterial peptides. (Methyl)lanthionines are posttranslationally introduced into the prepropeptides by biosynthetic enzymes that dehydrate serines and threonines and couple these dehydrated residues to cysteine residues. Thirty seven lantibiotic primary structures have been proposed to date, but little is known about the substrate specificity of the lantibiotic modifying enzymes. To define rules for the rational design of modified peptides, we compared all known lantibiotic structures by in silico analysis. Although no strict sequence motifs can be defined that govern the modification, statistical analysis demonstrates that dehydratable serines and threonines are more often flanked by hydrophobic than by hydrophilic amino acids. Serine residues escape dehydration more often than threonines. With these rules, novel hexapeptides were designed that either were predicted to become modified or will escape modification. The hexapeptides were fused to the nisin leader and expressed in a Lactococcus lactis strain containing the nisin modifying and export enzymes. The excreted peptides were analyzed by mass spectrometry. All designed fusion peptides were produced, and the presence or absence of modifications was found to be in full agreement with the predictions based on the statistical analysis. These findings demonstrate the feasibility of the rational design of a wide range of novel peptides with dehydrated amino acid residues.

Alanine↗
PubMed2006

Exchange of glutamine-217 to glutamate of Clostridium limosum exoenzyme C3 turns the asparagine-specific ADP-ribosyltransferase into an arginine-modifying enzyme.

C3-like ADP-ribosyltransferaseses are produced by Clostridium species, Bacillus cereus, and various Staphylococcus aureus strains. The exoenzymes modify the low-molecular-mass GTPases RhoA, B, and C. In structural studies of C3-like exoenzymes, an ARTT-motif (ADP-ribosylating turn-turn motif) was identified that appears to be involved in substrate specificity and recognition (Han, S., Arvai, A. S., Clancy, S. B., Tainer, J. A. (2001) J. Mol. Biol. 305, 95-107). Exchange of Gln217, which is a key residue of the ARTT-motif, to Glu in C3 from Clostridium limosum results in inhibition of ADP-ribosyltransferase activity toward RhoA. The mutant protein is still capable of NAD-binding and possesses NAD+ glycohydrolase activity. Whereas recombinant wild-type C3 modifies Rho proteins specifically at an asparagine residue (Asn41), Gln217Glu-C3 is capable of ADP-ribosylation of poly-arginine but not poly-asparagine. Soybean trypsin inhibitor, a model substrate for many arginine-specific ADP-ribosyltransferases, is modified by the Gln217Glu-C3 transferase. Also in C3 ADP-ribosyltransferases from Clostridium botulinum and B. cereus, the exchange of the equivalent Gln residue to Glu blocked asparagine modification of RhoA but elicited arginine-specific ADP-ribosylation. Moreover, the Gln217Glu-C3lim transferase was able to ADP-ribosylate recombinant wild-type C3lim at Arg86, resulting in decrease in ADP-ribosyltransferase activity of the wild-type enzyme. The data indicate that the exchange of one amino acid residue in the ARTT-motif turns the asparagine-modifying ADP-ribosyltransferases of the C3 family into arginine-ADP-ribosylating transferases.

ADP Ribose Transferases↗
PubMed1996

N (epsilon)-(carboxymethyl)lysine protein adduct is a major immunological epitope in proteins modified with advanced glycation end products of the Maillard reaction.

Long-term incubation of proteins with glucose leads to the formation of advanced glycation end products (AGE). Recent immunological studies have suggested the potential role of AGE in atherosclerosis, aging, and diabetic complications. We previously prepared a monoclonal (6D12) as well as a polyclonal anti-AGE antibody and proposed the presence of a common AGE structure(s) that may act as a major immunochemical epitope [Horiuchi, S., Araki, N., & Morino, Y. (1991) J. Biol. Chem. 266, 7329-7332]. The purpose of the present study was to determine the major epitope. Amino acid analysis of AGE-proteins indicated that N(epsilon)-(carboxymethyl)lysine (CML) was a major modified lysine residue. Immunologic studies demonstrated the positive reaction of 6D12 not only to all CML-modified proteins tested, but also to BSA modified with several aldehydes known to generate a CML-protein adduct, and a linear correlation between the CML contents of CML-BSA and their immunoreactivity to 6D12 up to approximately 8 mol/mol of protein. Further experiments with CML analogs revealed that the epitope of 6D12 is a CML-protein adduct with an important carbonyl group. In contrast to 6D12, our polyclonal anti-AGE antibody showed a significant but much weaker immunoreactivity to CML-BSA, suggesting that the polyclonal antibody contains two populations, one reactive to CML (CML-PA) and the other unreactive to CML (Non-CML-PA). Non-CML-PA separated from CML-PA by CML-BSA affinity chromatography did not react with all CML-modified preparations, but retained its property to react commonly with AGE preparations obtained from proteins, lysine derivatives, and monoaminocarboxylic acids. Therefore, it is clear that a CML-protein adduct is a major immunological epitope in AGE structures, but there still exist other major epitope(s) expressed commonly in AGE-proteins.

Animals↗
PubMed1998

What affects the effect of 2'-alkoxy modifications? 1. Stabilization effect of 2'-methoxy substitutions in uniformly modified DNA oligonucleotides.

The thermostability of hybrid duplexes with uniformly 2'-methoxy modified DNA strands (D'R and RD'), their unmodified DNA:RNA counterparts (DR and RD), and corresponded RNA:RNA (RR) duplexes for six sequences with different GC and deoxypyrimidine (dPy) content was measured. The linear correlation between the total stabilization effect of 2'-methoxy modifications (Delta DeltaG(o)37(D'R-DR)) and the relative stability of corresponding unmodified hybrids compared to the RR counterparts (Delta DeltaG(o)37(RR-DR)) suggests that the initial conformational and the thermodynamic state of the "parent" unmodified hybrid governs the effect of 2'-methoxy (and may be other 2'-alkoxy) modifications whose mechanism of action includes an S --> N conformational shift resulting in an RNA-like A-form duplex. We also found a correlation between the "hydrophobic" part of the total effect (Delta DeltaG(o)37(D'R-RR)) and the dA fraction in the modified DNA strand, suggesting that the "hydrophobic" effect of the 2'-methoxy groups results mainly from intraresidue steric effects increasing rigidity of the modified sugar rings. The correlations observed enabled us to predict the stability of hybrids with 2'-methoxy modified DNA strands for any sequence except for sequences with (dU)10 and (dA)10 strings.

Circular Dichroism↗
PubMed1998

Identification of the subunit and important target peptides of pig heart NAD-dependent isocitrate dehydrogenase modified by the affinity label adenosine 5'-O-[S-(4-bromo-2, 3-dioxobutyl)thiophosphate].

Pig heart NAD-dependent isocitrate dehydrogenase is inactivated by adenosine 5'-O-[S-(4-bromo-2,3-dioxobutyl)thiophosphate] (AMPS-BDB) with incorporation of 1.78 mol of reagent/mol of average subunit. Complete protection against the inactivation is provided by 20 mM isocitrate + 1 mM Mn2+, and the incorporation is decreased to about 1.3 mol of reagent/mol of average subunit. The addition of NAD, NADH, or Mn2+ alone has little effect on the functional changes produced by AMPS-BDB, while ADP gives only partial protection against the inactivation. The ability of ADP to decrease the Km for isocitrate is not affected by the AMPS-BDB modification of the enzyme. These results indicate that the isocitrate substrate site is the target of AMPS-BDB. The enzyme has three types of subunits with a tetramer having the composition alpha2 beta gamma. Here, [2-3H]AMPS-BDB-modified subunits are separated by HPLC on a C4 reverse-phase column, after the treatment of the modified enzyme with 4 M urea. The predominant radioactivity is distributed in alpha and gamma subunits. However, evidence based on recombination of subunits from modified and unmodified enzymes indicates that only labeling of the alpha subunit is responsible for inactivation by AMPS-BDB. Subsequently, the separated modified subunits were chemically cleaved by CNBr and then purified by HPLC using a C18 column. The labeled peptides were further digested by pepsin, purified by HPLC, and sequenced. These results indicate that R88 and R98 from the alpha subunit are the major targets of AMPS-BDB which cause inactivation and that these are at or near the isocitrate site of the enzyme.

Adenosine Diphosphate↗
PubMed1998

Characterization of serine and threonine phosphorylation sites in beta-elimination/ethanethiol addition-modified proteins by electrospray tandem mass spectrometry and database searching.

A new method for the characterization of serine and threonine phosphorylation sites in proteins has been developed. After modification of a phosphoprotein by beta-elimination/ethanethiol addition and conversion of phosphoserine and phosphothreonine residues to S-ethylcysteinyl or beta-methyl-S-ethylcysteinyl residues, the modified protein was subjected to proteolytic digestion. Resulting digests were analyzed by a combination of microbore liquid chromatography, electrospray ionization tandem (MS/MS) ion trap mass spectrometry and database searching to identify original phosphorylated residues. The computer program utilized (SEQUEST) is capable of identifying peptides and modified residues from uninterpreted MS/MS spectra, and using this method, all of the five known phosphorylation sites in bovine beta-casein were identified. Application of the method to multiply phosphorylated human high molecular weight neurofilament protein (NF-H) resulted in the identification of 21 peptides and their modified residues and hence, the in vivo phosphorylation sites. These included 26 KSP and 1 KTP site, all of which occur in the KSP repeat C-terminal tail domain (residues 502-823). One site at residue 518 was previously uncharacterized. A novel non-KSP serine at residue 421 near the KLLEGEE region in a IPFSLPE motif was characterized as phosphorylated (or glycosylated). The 27 characterized phosphorylation sites occur at S/TP residues in the following motifs: KSPVKEE, KSPAEAK, KSPEKEE, KSPAEVK, KSPEKAK, KSPPEAK, KSPVKAE, and KTPAKEE. On the basis of kinase consensus sequences, all of these motifs, including the previously unreported KTPAKEE motif, can be phosphorylated by proline-directed kinases. Advantages of the new method vis-a-vis our previously reported method [Jaffe, H., Veeranna, Shetty, K. T., and Pant, H. C. (1998) Biochemistry 37, 3931-3940] include (i) production of diastereomers eluting at different retention times increased the chances of peptide identification, (ii) increased hydrophobicity and hence retention time of the modified peptides, (iii) facilitation of positive ion production, and (iv) increased susceptibility to tryptic digestion as a result of conversion of negatively charged phosphorylated residues to neutral S-ethylcysteine or beta-methyl-S-ethylcysteine residues.

Amino Acid Sequence↗
PubMed1998

Dual recognition of double-stranded DNA by 2'-aminoethoxy-modified oligonucleotides: the solution structure of an intramolecular triplex obtained by NMR spectroscopy.

The solution structure of an intramolecular triple helical oligonucleotide has been solved by NMR. The third strand of the pyrimidine x purine x pyrimidine triplex is composed of 2'-aminoethoxy-modified riboses, whereas the remaining part of the nucleic acid is DNA. The structure around the aminoethoxy modification was obtained with the help of selective isotope labeling in conjunction with isotope-editing experiments. Dinucleotide steps and interstrand connectivities, as well as the complete backbone conformation of the triplex, were derived from J-couplings, NOEs, and 31P chemical shifts. The structure of this triplex, solved by distance geometry, explains the extraordinary stability and increase in rate of triplex formation induced by 2'-aminoethoxy-modified oligonucleotides: apart from the formation of seven base triples, a well-defined hydrogen-bonding network is formed across the Crick-Hoogsteen groove involving the amino protons of the aminoethoxy moieties and the phosphates of the purine strand of the DNA. The modified strand adopts a conformation which is close to an A-type helix, whereas the DNA duplex conformation is best described as an unwound B-type helix. The groove dimensions and helical parameters of the 2'-aminoethoxy-modified rY x dRdY triplex are surprisingly well conserved in comparison with DNA triplexes.

Computer Simulation↗
PubMed1999

Triplex formation of chemically modified homopyrimidine oligonucleotides: thermodynamic and kinetic studies.

We have investigated effects of chemical modifications of a third strand on the thermodynamic and kinetic properties of the triplex formation between a 23-bp duplex and each of four kinds of 15-mer chemically modified third strands using isothermal titration calorimetry and interaction analysis system. The chemical modifications of the third strand included one base modification, with replacement of thymine by uracil; two sugar moiety modifications, RNA and 2'-O-methyl-RNA; and one phosphate backbone modification, with replacement of phosphodiester by phosphorothioate backbone. The thermodynamic and kinetic parameters obtained were similar in magnitude at room temperature for the triplex formation with the base-modified and the sugar-modified third strands. By contrast, binding constant for the triplex formation with the third strand containing phosphorothioate backbone was much smaller by a factor of 10 than that for the other triplex formations. Kinetic analyses have also demonstrated that the third strand containing phosphorothioate backbone was much slower in the association step and much faster in the dissociation step than the other third strands, which resulted in the much smaller binding constant. The reason for the instability of the triplex with the third strand containing phosphorothioate backbone will be discussed. We conclude that, at least in the triplex formation with the chemically modified third strands studied in the present work, the modification of phosphate backbone of the third strand produces more significant effect on the triplex formation than the modifications of base and sugar moiety.

Base Sequence↗
PubMed2001

Thermo-responsive peptide-modified hydrogels for tissue regeneration.

Loosely cross-linked hydrogels consisting of N-isopropylacrylamide (NIPAAm) and acrylic acid (AAc) were synthesized, characterized, and used as model scaffolds for studying cell-material interactions in three-dimensions (3D). The AAc groups were functionalized with peptides containing the -RGD- and -FHRRIKA- sequences found in bone sialoprotein. Chemical modification of the hydrogels was verified via solid-state (1)H nuclear magnetic resonance spectroscopy, lower critical solution temperature studies, and volume change studies. The peptide-modified hydrogels were pliable at 22 degrees C and could be injected through a small-diameter aperture. Rat calvarial osteoblasts (RCO) seeded into the peptide-modified hydrogels were viable for at least 21 days of in vitro culture. The RCO spread more and demonstrated significantly greater proliferation when cultured within the peptide-modified hydrogels, as compared to control hydrogels. These peptide-modified P(NIPAAm-co-AAc) hydrogels serve as useful tools for studying cell-material interactions within 3D structures and have the potential to be used as injectable scaffolds for tissue engineering applications.

Acrylamides↗
PubMedDate not supplied

Hydroxyapatite surface modified by L-lactic acid and its subsequent grafting polymerization of L-lactide.

A new method of surface modification of hydroxyapatite nanoparticles (n-HA) by surface grafting reaction of l-lactic acid and ring-opening polymerization of l-lactide (LLA) was developed. Two modified HA nanoparticles were obtained: HA modified by l-lactic acid (l-HA) and HA grafting with poly(l-lactide) (PLLA; p-HA). The modified surface of n-HA was attested by Fourier transformation infrared, (31)P MAS NMR, and thermal gravimetric analysis. The results showed that l-lactic acid could be easily grafted onto the n-HA surface by forming a Ca carboxylate bond and initiated by the hydroxyl group of the grafted l-lactic acid and LLA could be graft-polymerized onto the n-HA surface in the presence of stannous octanoate. The highest grafting amounts of l-lactic acid and PLLA were about 33 and 22 wt %, respectively. The modified HA/PLLA composites showed good mechanical properties and uniform microstructure. The tensile strength and modulus of the p-HA/PLLA composite containing 15 wt % of p-HA were 67 MPa and 2.1 GPa, respectively, while those of the n-HA/PLLA composites were 45 MPa and 1.7 GPa, respectively. The elongation at the break of the l-HA/PLLA composite containing 15 wt % l-HA could reach 44%, in comparison with 6.5% of the n-HA/PLLA composites containing 15 wt % n-HA.

Durapatite↗
PubMedDate not supplied

Modified electrodes with synthetic biocatalytic membranes.

A biocatalytic membrane based on an immobilized enzyme molecule has been prepared. Oxidative electropolymerization of 8-hydroxyquinaldine (8-OHQ) monomers in 0.2 M, pH 7 phosphate buffer containing glucose oxidase (GOx) has been carried out to modify the surfaces of GC, Au, and Pt rotating disk electrodes. The biocatalytic properties of the synthesized membrane were characterized by studying the catalytic activities of the immobilized GOx. Signals obtained from modified GC electrodes with this biomembrane were mainly attributed to the immobilized GOx. Signals obtained from modified Pt or Au electrodes were due to the combined contribution of the enzyme and the native electrode's material. The potential analytical applications of these modified electrodes as bioelectrochemical sensors were also investigated.

Biotechnology↗
PubMedDate not supplied

Selection of modified oligonucleotides with increased target affinity via MALDI-monitored nuclease survival assays.

Reported here is how modified oligonucleotides with increased affinity for DNA or RNA target strands can be selected from small combinatorial libraries via spectrometrically monitored selection experiments (SMOSE). The extent to which target strands retard the degradation of 5'-acyl-, 5'-aminoacyl-, and 5'-dipeptidyl-oligodeoxyribonucleotides by phosphodiesterase I (EC 3.1.4.1) was measured via quantitative MALDI-TOF mass spectrometry. Oligonucleotide hybrids were prepared on solid support, and nuclease selections were performed with up to 10 modified oligonucleotides in one solution. The mass spectrometrically monitored experiments required between 120 and 300 pmol of each modified oligonucleotide, depending on whether HPLC-purified or crude compounds were employed. Data acquisition and analysis were optimized to proceed in semiautomated fashion, and functions correcting for incomplete degradation during the monitoring time were developed. Integration of the degradation kinetics provided "protection factors" that correlate well with melting points obtained with traditional UV melting curves employing single, pure compounds. Among the components of the five libraries tested, three were found to contain 5'-substituents that strongly stabilize Watson--Crick duplexes. Selecting and optimizing modified oligonucleotides via monitored nuclease assays may offer a more efficient way to search for new antisense agents, hybridization probes, and biochemical tools.

Automation↗
PubMedDate not supplied

A mass spectrometry screening method for antiaggregatory activity of proteins covalently modified by combinatorial library members: application to sickle hemoglobin.

A homogeneous assay, based on electrospray mass spectrometry, is described for identifying compounds in a combinatorial library that covalently modify a protein and thereby enhance its solubility. The technique is based on measuring the distribution of modified proteins in the supernatant versus aggregate. Compounds having the greatest anti-aggregatory activity are those with the highest supernatant/aggregate ratio. Mass is used as a marker to identify which covalent modifier in the library is involved. An exploratory study is presented which demonstrates that the antisickling activity of a family of isothiocyanates, as measured by the standard C(sat) assay, correlates well (r(2) = 0.98) with the mass spectrometry analysis of the supernatant/aggregate distribution. The technique has potential for screening libraries capable of covalently modifying other proteins of clinical interest, e.g., Alzheimer's, Huntington's, and various prion related diseases.

Antisickling Agents↗
PubMed2004

Identification of the reactive oxygen species responsible for carbon tetrachloride degradation in modified Fenton's systems.

The reactive oxygen species responsible for the transformation of carbon tetrachloride (tetrachloromethane, CT) by modified Fenton's reagent using hydrogen peroxide (H2O2) concentrations >0.1 M was investigated. Addition of the hydroxyl radical scavenger 2-propanol to modified Fenton's reactions did not significantly lower CT transformation rates. Scavenging by 2-propanol not only confirmed that hydroxyl radicals are not responsible for CT destruction, but also suggested that a major product of an iron (III)-driven initiation reaction, superoxide radical anion (O2-), is the species responsible for CT transformation. To investigate this hypothesis, CT degradation was studied in aqueous KO2 reactions. Minimal CT degradation was found in CT-KO2 reactions; however, when H2O2 was added to the KO2 reactions at concentrations similar to those in the modified Fenton's reactions (0.1, 0.5, and 1 M), CT degradation increased significantly. Similar results were obtained when 1 M concentrations of other solvents were added to aqueous KO2 reactions, and the observed first-order rate constant for CT degradation correlated strongly (R2 = 0.986) with the empirical solvent polarity (E(T)N) of the added solvents. The results indicate that even dilute concentrations of solvents, including H202, can increase the reactivity of O2- in water, probably by changing its solvation sphere. The higher reactivity of O2- generated in modified Fenton's reagent, which has a less polar nature due to the presence of H2O2, may result in a wider range of contaminant degradation than previously thought possible.

Carbon Tetrachloride↗
PubMed1999

Electrochemical Detection of Single-Stranded DNA Using Polymer-Modified Electrodes.

Glassy carbon electrodes modified by reductive electropolymerization of a thin film of poly[Ru(vbpy)(3)(2+)] or poly[Ru(vbpy)(3)(2+)/vba] (vbpy = 4-vinyl-4'-methyl-2,2'-bipyridine and vba = p-vinylbenzoic acid) were prepared. The Ru(III/II) couples for the polymer films were reversible in nonaqueous solution but were irreversible in aqueous media. The films modified with poly[Ru(vbpy)(3)(2+)] catalyzed the oxidation of aqueous guanosine 5'-monophosphate (GMP) and poly[G], producing a current enhancement in the Ru(III/II) couple for the polymer film. The catalysis was due both to electrostatic condensation of GMP and poly[G] to the Ru-modified surface and to more facile electron transfer to the Ru(III) centers in the polymer compared to the bare electrode. The presence of GMP in solution decreased the extent of decomposition of Ru(III). When single-stranded DNA containing multiple guanines was attached to the electrode modified with the poly[Ru(vbpy)(3)(2+)/vba] copolymer, enhancement of 8-13 &mgr;A for the Ru(III/II) couple was observed with 8 pmol of attached DNA. This degree of enhancement corresponds to a current efficiency of 65% based on a one-electron oxidation of guanine.

Journal Article↗
PubMed2003

Oligo-2'-deoxyribonucleotides containing uracil modified at the 5-position with linkers ending with guanidinium groups.

We report here the synthesis and binding studies of oligo-2'-deoxyribonucleotides (ODNs) containing 2'-deoxyuridines, modified at the 5-position by linkers ending with either one or two guanidinium groups. Comparison was made with ODNs containing 2'-deoxyuridines modified at the 5-position with linkers ending with either two or one amino groups. One or two modified 2'-deoxyuridines were incorporated into pyrimidine strands, and their influence on the stability of duplex (with both DNA and RNA targets) and triplex structures was studied. The strongest stabilization was obtained with modified ODNs containing guanidinium groups. This result confirms that the reduction of the global negative charge number on one strand is an important parameter in the stability of duplex and triplex structures.

Cross-Linking Reagents↗
PubMed2003

Interaction between ketopantolactone and chirally modified Pt investigated by attenuated total reflection IR concentration modulation spectroscopy.

The combination of ATR-IR and modulation spectroscopy allowed for the study of the interaction of ketopantolactone with Pt/Al2O3 films chirally modified by cinchonidine under hydrogenation conditions. The spectra reveal a significant influence of ketopantolactone on the adsorption of the modifier and indicate a N-H-O hydrogen bond between modifier and reactant. The latter was corroborated by a comparative study with N-methyl cinchonidine chloride modified Pt/Al2O3.

4-Butyrolactone↗