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At least 325 records · Page 18Linked to original sources
PubMed1998

Valine substituted winter flounder 'antifreeze': preservation of ice growth hysteresis.

Three mutant polypeptides of the type I 37-residue winter flounder 'antifreeze' protein have been synthesized. All four threonine residues in the native peptide were been mutated to serine, valine and glycine respectively and two additional salt bridges were incorporated into the sequences in order to improve aqueous solubility. The peptides were analyzed by nanoliter osmometry, the 'ice hemisphere' test, the 'crystal habit' test, measurement of ice growth hysteresis and CD spectroscopy. While the valine and serine mutants retain the alpha-helical structure, only the valine mutant retains 'antifreeze' activity similar to that of the native protein. These data show that the threonine hydroxyl groups do not play a crucial role in the accumulation of the native 'antifreeze' protein at the ice/water interface and the inhibition of ice growth below the equilibrium melting temperature.

Amino Acid Substitution↗
PubMed2003

Chiral separation of underivatized and dansyl amino acids by ligand-exchange micellar electrokinetic capillary chromatography using a copper(II)-L-valine complex as selector.

Ligand-exchange micellar electrokinetic capillary chromatography was used for the chiral resolution of underivatized and dansyl amino acid enantiomers simultaneously. The separation was achieved by chiral copper(II)-L-valine complexes incorporated in micelles of sodium dodecyl sulfate (SDS). The enantioresolution was strongly affected by SDS and a concentration of 20 mM SDS was shown to be necessary for the separation. Other impacting factors were investigated including pH, the molar ratio of copper(II) to L-valine and the total concentration of complex. Using the proposed method, 11 different dansyl amino acids and two underivatized amino acids were separated successfully with a running electrolyte of 20 mM NH4OAc, 4 mM CuSO4, 8 mM L-valine and 20 mM SDS at pH 9.0 in less than 25 min. Experiments were also performed with other amino acid ligands in order to vary the stability and the sterical arrangement of the copper(II) complexes and the possible chiral recognition mechanism was also discussed briefly.

Amino Acids↗
PubMed2001

In vitro activity of MEKK2 and MEKK3 in detergents is a function of a valine to serine difference in the catalytic domain.

MEKK2 and MEKK3 are mitogen-activated protein kinase kinase kinases (MAP3 kinases) of 70 and 71 kDa respectively that are markedly homologous (94%) in their kinase domains. Both MEKK2 and MEKK3 are able to activate the Jun kinase pathway in vivo. However, following routine immunoprecipitation in Triton X-100, MEKK2 but not MEKK3 is able to effectively phosphorylate both SEK-1 and MEK-1 and to undergo autophosphorylation. Unexpectedly, both MEKK2 and MEKK3 are functional in an in vitro kinase assay when cells are solubilized with the closely related detergent, NP-40. Given the high homology between these kinases, we set out to relate this differential sensitivity to Triton X-100 to differences in primary structure. A set of chimeric molecules were generated and the loss of activity in Triton X-100 mapped to kinase domain II/III and specifically to serine 390 of MEKK3 and valine 384 of MEKK2, residues immediately N-terminal to the active site lysine. Mutation of serine 390 of MEKK3 to a valine (as is found in MEKK2) conferred catalytic activity to MEKK3 in Triton X-100 whereas the reciprocal alteration of valine 384 of MEKK2 to a serine conferred lack of activity in Triton X-100 to MEKK2. Search of the protein database identified only three kinases, MEKK3, Pbs2p and Dd-PKI, with a serine or threonine at this site. The presence of a serine or threonine adjacent to the active site lysine in protein kinases is rare and, in MEKK3, results in detergent instability.

Amino Acid Sequence↗
PubMed1985

Mineral nutrition of Aerobacter aerogenes for valine production in a synthetic medium.

The effect of a number of mineral salts, like dipotassium hydrogen phosphate, magnesium sulphate, and sodium chloride, and of some trace elements including iron, copper, cobalt, nickel, molybdenum, and calcium, on the production of valine by Aerobacter aerogenes in a synthetic medium was investigated. It was found that all the mineral salts were necessary for valine formation. Among the trace elements, iron and molybdenum were found to be necessary in minute concentrations for the optimum yield of the amino acid, while all the others had an adverse effect on valine production, even at lower levels.

Culture Media↗
PubMed1999

Determination of extracellular and intracellular enrichments of [1-(13)C]-alpha-ketoisovalerate using enzymatically converted [1-(13)C]-valine standardization curves and gas chromatography-mass spectrometry.

We report a validated method for the determination of extra- and intracellular [1-(13)C]-alpha-ketoisovalerate ([1-(13)C]-KIV) enrichments by gas chromatography-mass spectrometry. Standardization curves were prepared by enzymatic oxidation of [1-(13)C]-valine enriched standards of known composition. Slopes of [1-(13)C]-valine standardization curves (mean+/-SD: 0.99+/-0.02, n=5) and [1-(13)C]-KIV standardization curves (mean+/-SD: 0.98+/-0.01, n=7) were not significantly different. The method was applied for the determination of [1-(13)C]-KIV enrichments in plasma and tissues during [1-(13)C]-valine infusion in a piglet. [1-(13)C]-KIV enrichment could be determined+/-0.1 MPE (C.V. 1%), and extracellular [1-(13)C]-KIV enrichment was a reliable estimate of intracellular (skeletal muscle, bone growth plate) [1-(13)C]-KIV enrichment.

Animals↗
PubMed1994

Conformation of valine side chains in ribonuclease T1 determined by NMR studies of homonuclear and heteronuclear 3J coupling constants.

A conformational analysis of the valine side chains of ribonuclease T1 (RNase T1) was performed using NMR spectroscopy, in particular homonuclear (1H, 1H and 13C, 13C) and heteronuclear (1H, 15N and 1H, 13C) vicinal spin-spin coupling constants as obtained from E.COSY-type NMR experiments. The coupling constants related to the chi 1 dihedral angle in valine, 3JH alpha H beta, 3JNH beta, 3JC'H beta, 3JH alpha C gamma 1, 3JH alpha C gamma 2, 3JC'C gamma 1, and 3JC'C gamma 2, were evaluated in a quantitative manner. The analysis of 3J data allowed for the stereospecific assignment of the valine methyl resonances. On the basis of various models for motional averaging of coupling constants, a fit of the torsion angles chi 1 to a set of the experimental 3J coupling constants (3JH alpha H beta, 3JNH beta, 3JC'H beta) was carried out. The resulting side-chain conformations were examined with respect to NOE distance informations. Single rotameric states emerged for Val16, Val67, Val79, and Val101, while conformational equilibria between staggered rotamers were found for Val33 and Val78. Using a different model approach, Val52 and Val89 are also likely to exhibit unimodal chi 1 angle distributions. The analysis was found to depend critically on the set of Karplus parameters used. Except for Val52 and Val78, the predominant rotamers obtained from 3J coupling informations agree with the conformation in the crystal structure of ribonuclease T1 (Martinez-Oyanedel et al., 1991).

Magnetic Resonance Spectroscopy↗
PubMed1990

Use of site-directed mutagenesis to identify valine-573 in the S'1 binding site of rat neutral endopeptidase 24.11 (enkephalinase).

On the basis of the identity of a segment of the amino acid sequence within the active site of the bacterial enzyme thermolysin and the mammalian enzyme neutral endopeptidase 24.11, the possible involvement of valine-573 of neutral endopeptidase 24.11 in substrate binding was investigated. Valine-573 was changed to leucine and to alanine by site-directed mutagenesis. The effect of these mutations on inhibitor binding and substrate catalysis was examined with a series of compounds containing variable P'1 residues. With a small P'1 residue such as alanine, both mutant enzymes exhibited kinetic properties essentially the same as the wild-type enzyme. However, with larger P'1 residues such as phenylalanine, tyrosine, and leucine, the Val573Leu mutant showed a 24-100-fold decrease in inhibitor affinity. Similarly substrates containing bulky P'1 residues showed a 10-40-fold decrease in Vmax with little change in Km. In contrast, the Val573Ala mutant showed only modest changes in terms of inhibitor binding or substrate turnover. These results support the proposed role of valine-573 as a part of the hydrophobic binding pocket, S'1 binding subsite, of neutral endopeptidase 24.11.

Amino Acid Sequence↗
PubMed1977

Editing mechanisms in protein synthesis. Rejection of valine by the isoleucyl-tRNA synthetase.

Although the isoleucyl-tRNA synthetase from Escherichia coli (IRS) does not catalyze the overall mischarging of tRNAIle with valine, it does undergo the first step of the reaction, the formation of an IRS-Val-AMP complex. The addition of tRNAIle to this complex leads to its quantitative hydrolysis and the IRS acts as an ATP pyrophosphate in the presence of valine and tRNAIle (Baldwin, A.N., and Berg, P. (1966), J. Biol. Chem. 241, 839). It is shown that during the ATP pyrophosphatase reaction: (a) IRS forms an IRS-Val-AMP complex; (b) the turnover number of the ATP pyrophosphatase reaction is the same at the rate constant for the transfer of isoleucine from IRS-Ile-AMP to tRNAIle over a wide range of temperature and pH; (c) mischarged Val-tRNAIle is hydrolyzed by IRS with a turnover number of 10 s-1 at pH 7.78 and 25 degrees C, compared with a value of 1.2 s-1 for the transfer of isoleucine from IRS-Ile-AMP to tRNA or for the ATP pyrophosphatase reaction. Although this appears to be consistent with an editing mechanism in which there is a slow transfer of the valine from the IRS-Val-AMP to tRNAIle follwed by the rapid hydrolytic step, as recently found for the rejection of threonine by the valyl-tRNA synthetase, there is an inconsistency. This scheme predicts that on mixing IRS.[14C]Val-AMP with tRNAIle there should be a transient misacylation of the tRNA such that about 10% of the [14C]Val is present as [14C]Val-tRNAIle at the peak. But 0.8% or less is found. This could possibly be caused by the IRS having a higher hydrolytic activity during the mischarging reaction than is measured on mixing the unligated enzyme with performed Val-tRNAIle. Alternatively, a two-stage editing mechanism must be considered in which the majority of the Val-AMP is destroyed before the transfer to tRNA in the major editing step, while the hydrolytic activity of the IRS towards Val-tRNAIle is a second editing step to mop up any mischarged tRNA formed by the Val-AMP escaping the first editing step. It is shown that the "kinetic proofreading" mechanism of Hopfield is not consistent with the experimental data.

Amino Acyl-tRNA Synthetases↗
PubMed1996

Physical and biological characteristics of the antitumor drug actinomycin D analogues derivatized at N-methyl-L-valine residues.

The crystal structure of the DNA-actinomycin D (AMD) complex and a simple molecular modeling study indicated that AMD analogues derivatized at N-methyl-L-valine residues (fifth amino acid residue in the cyclic depsipeptide of AMD) could bind to DNA as strongly as the parent AMD. The analogues in which N-methyl-L-valine residues were replaced with L- and D-forms of N-methylvalines, N-methylthreonines, N-methylphenylalanies, N-methyltyrosines, and N-methyl-O-methyltyrosines have been totally synthesized. The characteristics of binding of the analogues to various DNAs including DNA-1 [d(TATATATGCATATATA)], DNA-2 [d(TATATACGCGTATATA)], DNA-3 [d(ATATATAGCTATATAT)], and DNA-4 [d(ATATATGGCCATATAT)] have been examined by using visible absorption spectrum methods. The association constants calculated from the absorption spectra indicate that the modifications of the N-methyl-L-valine residues in the AMD molecule do affect the DNA binding characteristics of the analogues. The L-aromatic analogues bind slightly better than the L-aliphatic analogues except for binding to DNA-1 (-TGCA-), whereas the D-aliphatic analogues bind consistently better than the D-aromatic analogues. In the L-form analogues, the L-Tyr analogue has the highest overall association constant, whereas the D-Val analogue has the highest association constant among the D-form analogues. In spite of substitution of bulky aromatic groups, the D-aromatic analogues bind to the DNA-1 quite well. However, D-aromatic analogues have significantly reduced their binding capacities to the other DNAs, indicating that the substitution of the D-aromatic residues creates a unique four-base sequence preference (-TGCA-). The RNA polymerase inhibitory activities of the AMD analogues in vivo have been examined using human cells (HeLa). All AMD analogues except for the L-Thr analogues severely inhibit RNA synthesis at relatively low drug concentrations. The D-Val, L-OMT, L-Phe, and D-Phe analogues inhibit RNA synthesis more strongly than the natural antibiotic (AMD itself).

Antibiotics, Antineoplastic↗
PubMed1996

Monitoring of in vitro and in vivo exposure to sulfur mustard by GC/MS determination of the N-terminal valine adduct in hemoglobin after a modified Edman degradation.

We report that exposure to the chemical warfare agent sulfur mustard can be monitored by means of a modified Edman degradation involving selective release of the N-terminal valine adduct of hemoglobin with the agent. The degree of alkylation of the N-terminal valine in human hemoglobin is approximately 1-2% of the total alkylation induced in hemoglobin upon treatment of human blood with sulfur mustard. After modified Edman degradation, followed by derivatization with heptafluorobutyric anhydride, the obtained pentafluorophenyl thiohydantion derivative of the valine adduct could be analyzed at a > or = 0.5 fmol level by means of GC/MS under negative ion chemical ionization conditions. Applying this procedure, in vitro exposure of human blood to > or = 0.1 microM of sulfur mustard could be determined. In vivo exposure of guinea pigs could also be established at 48 h after intoxication intravenously with 0.5 mg/kg (0.06 LD50) of the agent.

Alkylation↗
PubMed2001

Valine 77 of heterocystous ferredoxin FdxH2 in Anabaena variabilis strain ATCC 29413 is critical for its oxygen sensitivity.

Ferredoxins are small iron sulfur proteins necessary for electron donation. FdxH1 and FdxH2 are associated with two different nif gene clusters where they transfer electrons for the reduction of nitrogenase complex. FdxH1 was observed to be stable towards oxygen, whereas, FdxH2 was relatively unstable. We had identified the amino acid involved in oxygen sensitivity of ferredoxin protein using protein modeling. The exchange of valine to leucine at position 77 was critical for ferredoxin proteins in relation to its oxygen sensitivity. This exchange leads to a longer side chain, which inhibits the accessibility of oxygen to the iron sulfur cluster. Site directed mutagenesis and in vitro experiments confirms that valine indeed is involved in the oxygen sensitivity. The exchange of leucine to valine in FdxH1 makes it oxygen unstable. Thus, from the above results we can conclude that the position of leucine at position 77 is critical for oxygen sensitivity of ferredoxin and protein modeling can be used to identify specific amino acids in other oxygen-sensitive proteins.

Amino Acid Substitution↗
PubMed2004

Disease associated mutations at valine 804 in the RET receptor tyrosine kinase confer resistance to selective kinase inhibitors.

We have recently demonstrated that the pyrazolopyrimidines PP1 and PP2 and the 4-anilinoquinazoline ZD6474 display a strong inhibitory activity (IC(50)< or =100 nM) towards constitutively active oncogenic RET kinases. Here, we show that most oncogenic MEN2-associated RET kinase mutants are highly susceptible to PP1, PP2 and ZD6474 inhibition. In contrast, MEN2-associated swap of bulky hydrophobic leucine or methionine residues for valine 804 in the RET kinase domain causes resistance to the three compounds. Substitution of valine 804 with the small amino- acid glycine renders the RET kinase even more susceptible to inhibition (ZD6474 IC(50): 20 nM) than the wild-type kinase. Our data identify valine 804 of RET as a structural determinant mediating resistance to pyrazolopyrimidines and 4-anilinoquinazolines.

Carcinoma, Medullary↗
PubMed1970

The effect of electrical stimulation on the incorporation of L-[U-14C] valine into the protein of chopped tissue from guinea-pig cerebral cortex.

1. Chopped tissue from guinea-pig cerebral cortex carried out an energy-dependent incorporation of [(14)C]valine into protein. 2. At all times studied the nuclear fraction of the homogenized tissue accounted for about 25% of the total labelled protein. 3. Electrical stimulation at first increased, but subsequently decreased, the rate of incorporation of [(14)C]valine into protein of the chopped tissue. 4. The initial increase in the incorporation of [(14)C]valine into protein occurred in the nuclear fraction. At later times electrical stimulation decreased the incorporation into all the subcellular fractions, but the relative contribution of the nuclear fraction to the total labelled protein increased. 5. These changes are discussed in relation to the changes in the rates of respiration, glycolysis, high-energy phosphate content and intracellular Na(+) and K(+) concentrations, which were measured under the same experimental conditions as those used to study protein synthesis.

Adenosine Triphosphate↗
PubMed1984

Stereochemistry of the incorporation of valine methyl groups into methylene groups in cephalosporin C.

'Chiral methyl valines', i.e. samples of valine labelled stereospecifically in the methyl groups with 2H and 3H, were incorporated into cephalosporin C by a suspension of washed cells of Cephalosporium acremonium. Analysis by 3H n.m.r. of the cephalosporin C produced showed that the conversion of the 3-pro-S-methyl group of valine into the acetoxymethyl side-chain was a highly stereospecific process. By contrast, conversion of the 3-pro-R-methyl group into the endocyclic methylene group of the dihydrothiazine ring was shown to proceed by a non-stereospecific process.

Acremonium↗
PubMed1994

Substrate specificity of L-delta-(alpha-aminoadipoyl)-L-cysteinyl-D-valine synthetase from Cephalosporium acremonium: demonstration of the structure of several unnatural tripeptide products.

Potential substrates for L-delta-(alpha-aminoadipoyl)-L-(cysteinyl)-D-valine (ACV) synthetase were initially identified using both the amino-acid-dependent ATP<-->pyrophosphate exchange reaction catalysed by the enzyme and the incorporation of 14C-radiolabelled cysteine and valine into potential peptide products. S-Carboxymethylcysteine was an effective substitute for alpha-aminoadipate and both allylglycine and vinylglycine could substitute for cysteine, indicating that the thiol group of cysteine is not essential for peptide formation. L-allo-Isoleucine but not L-isoleucine substituted effectively for valine. The structures of the presumed peptide products derived from these amino acids were confirmed by combined use of electrospray-ionization m.s. (e.s.m.s.) and 1H n.m.r. These results clearly indicate that, in common with other peptide synthetases, but in contrast with ribosomal peptide synthesis, ACV synthetase has a relatively broad substrate specificity.

2-Aminoadipic Acid↗
PubMed1997

Purification and characterization of delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase from Penicillium chrysogenum.

delta-(L-alpha-Aminoadipyl)-L-cysteinyl-D-valine synthetase (ACVS) from Penicillium chrysogenum was purified to homogeneity by a combination of (NH4)2SO4 precipitation, protamine sulphate treatment, ion-exchange chromatography, gel filtration and hydrophobic interaction chromatography. The molecular mass of ACVS was estimated with native gradient gel electrophoresis and SDS/PAGE. The native enzyme consisted of a single polymer chain with an estimated molecular mass of 470 kDa. The denatured enzyme had an estimated molecular mass of 440 kDa. The influence of different reaction parameters such as substrates, cofactors and pH on the activity of the purified ACVS was investigated. The Km values for the three precursor substrates L-alpha-aminoadipic acid, L-cysteine and L-valine were determined as 45, 80 and 80 microM respectively, and the optimal assay concentration of ATP was found to be 5 mM (with 20 mM MgCl2). The dimer of the reaction product bis-delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (bisACV) gave feedback inhibition of the purified ACVS; the inhibition parameter KbisACV was determined as 1.4 mM. Furthermore dithiothreitol was shown to inhibit the purified ACVS. From the addition of a glucose pulse to a steady-state glucose-limited continuous culture of P. chrysogenum it was found that there is glucose repression of the synthesis of ACVS and that there must be a constant turnover of ACVS owing to synthesis and degradation.

2-Aminoadipic Acid↗
PubMed2000

Clinicopathological phenotype of codon 129 valine homozygote sporadic Creutzfeldt-Jakob disease.

The naturally occurring polymorphism at codon 129 of the human prion protein gene (PRNP) influences susceptibility to sporadic Creutzfeldt-Jakob Disease (CJD); the majority of the patients are methionine homozygotes at this locus, while valine homozygotes represent only 10% of cases. The aim was to study the clinical and neuropathological phenotype of sporadic CJD in valine homozygotes, to estimate the reliability of current clinical diagnostic criteria, and to identify any consistent and distinct features. Twelve cases of sporadic CJD with a codon 129 valine homozygote genotype were identified at the National CJD Surveillance Unit in Edinburgh. In addition to a retrospective clinical analysis, tissue blocks were stained by conventional techniques and by immunocytochemistry for prion protein. Frozen brain tissue was available from five cases for Western blot analysis of PrPRES, which in all cases showed a type 2 mobility. The cases included four males and eight females, average age 63.6 years, with a mean duration of illness of 6 months. Eleven patients presented with ataxia, and none had the characteristic EEG changes found in sporadic CJD. The neuropathological phenotype comprised spongiform change and prion protein immunopositivity most marked in the subcortical grey matter and cerebellum, prion protein positive plaque-like deposits in all regions, laminar deposition of prion protein in the cerebral cortex, and hippocampal involvement (which is seldom reported in sporadic CJD). In conclusion, these cases exhibited a fairly uniform phenotype, which is relatively distinct from sporadic CJD in methionine homozygotes, and thus diagnosis may be difficult using existing clinical criteria.

Aged↗
PubMed2002

Valine-alanine manganese superoxide dismutase polymorphism is not associated with alcohol-induced oxidative stress or liver fibrosis.

The role of genetic factors in the pathogenesis of alcohol-induced liver disease (ALD) is receiving increasing attention. Recently, it has been reported that homozygosity for a valine to alanine substitution in the mitochondrial targeting sequence of manganese superoxide dismutase (Mn-SOD) represents a risk factor for severe ALD. Because this mutation is postulated to modify enzyme transport into mitochondria, we have sought confirmatory evidence of this association in a larger group of patients and investigated whether this polymorphism might influence alcohol-induced oxidative stress. Genotyping for the valine-alanine (Val-Ala) polymorphism of the Mn-SOD gene in 281 patients with advanced ALD (cirrhosis/fibrosis) and 218 drinkers without liver disease showed no differences in either the heterozygote (55% vs. 50%) or the homozygote (19% vs. 23%) frequency for the alanine allele. By measuring the titers of circulating antibodies against oxidized cardiolipin (OX-CL) and malondialdehyde (MDA) or hydroxy-ethyl radical (HER) adducts as markers of oxidative stress, we found a significant increase in ALD patients compared with healthy controls. However, the carriers of the alanine Mn-SOD allele had titers of anti-MDA, anti-HER, and anti-OX-CL IgG comparable with heterozygotes and patients homozygous for the valine allele. Similarly, the frequency of subjects with antibody titers above the 95th percentile of controls was not increased among homozygotes for the alanine Mn-SOD allele. In conclusion, in our population Val-Ala polymorphism in Mn-SOD influences neither susceptibility to alcohol-induced liver fibrosis nor alcohol-induced oxidative stress.

Alanine↗