Identification of valine/leucine/isoleucine and threonine/alanine/glycine proton-spin systems of Escherichia coli adenylate kinase by selective deuteration and selective protonation.
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ABSTRACT:
SUBMITTER: Bock-Mobius I
PROVIDER: S-EPMC1150215 | biostudies-other | 1991 Jan
REPOSITORIES: biostudies-other
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Project description:The aliphatic hydrophobic amino acid residues-alanine, isoleucine, leucine, proline and valine-are among the most common found in proteins. Their structural role in proteins is seemingly obvious: engage in hydrophobic interactions to stabilize secondary, and to a lesser extent, tertiary and quaternary structure. However, favorable hydrophobic interactions involving the sidechains of these residue types are generally less significant than the unfavorable set arising from interactions with polar atoms. Importantly, the constellation of interactions between residue sidechains and their environments can be recorded as three-dimensional maps that, in turn, can be clustered. The clustered average map sets compose a library of interaction profiles encoding interaction strengths, interaction types and the optimal 3D position for the interacting partners. This library is backbone angle-dependent and suggests solvent and lipid accessibility for each unique interaction profile. In this work, in addition to analysis of soluble proteins, a large set of membrane proteins that contained optimized artificial lipids were evaluated by parsing the structures into three distinct components: soluble extramembrane domain, lipid facing transmembrane domain, core transmembrane domain. The aliphatic residues were extracted from each of these sets and passed through our calculation protocol. Notable observations include: the roles of aliphatic residues in soluble proteins and in the membrane protein's soluble domains are nearly identical, although the latter are slightly more solvent accessible; by comparing maps calculated with sidechain-lipid interactions to maps ignoring those interactions, the potential extent of residue-lipid and residue-interactions can be assessed and likely exploited in structure prediction and modeling; amongst these residue types, the levels of lipid engagement show isoleucine as the most engaged, while the other residues are largely interacting with neighboring helical residues.
| S-EPMC10086146 | biostudies-literature
Project description:1. The reciprocal interference between l-leucine, l-isoleucine and l-valine during absorption was studied in rats both in vivo and with an everted-sac preparation in vitro. 2. After feeding with the amino acids alone there was a considerable increase in their concentration in the intestinal lumen followed by a rapid disappearance, indicating efficient absorption. Absorption was reflected by a high concentration of the respective amino acids in the portal plasma. Isoleucine and valine inhibited the absorption of leucine, and leucine inhibited the absorption of isoleucine and valine. Inhibition of absorption by the interfering amino acid was generally partly overcome after 30-60min., probably through the absorption of the interfering amino acid. At that time the rise in the concentration of the amino acid in portal plasma began. 3. These results were confirmed by experiments in vitro: isoleucine and valine inhibited the absorption rate of leucine, and leucine that of isoleucine and valine. 4. Active absorption of amino acids was rapid at low concentrations and depressed at higher concentrations.
| S-EPMC1265085 | biostudies-other
Project description:A novel fluorescence sensing system for branched-chain amino acids (BCAAs) was developed based on engineered leucine/isoleucine/valine-binding proteins (LIVBPs) conjugated with environmentally sensitive fluorescence probes. LIVBP was cloned from Escherichia coli and Gln149Cys, Gly227Cys, and Gln254Cys mutants were generated by genetic engineering. The mutant LIVBPs were then modified with environmentally sensitive fluorophores. Based on the fluorescence intensity change observed upon the binding of the ligands, the MIANS-conjugated Gln149Cys mutant (Gln149Cys-M) showed the highest and most sensitive response. The BCAAs Leu, Ile, and Val can each be monitored at the sub-micromolar level using Gln149Cys-M. Measurements were also carried out on a mixture of BCAFAs and revealed that Gln149Cys-M-based measurement is not significantly affected by the change in the molar ratio of Leu, Ile and Val in the sample. Its high sensitivity and group-specific molecular recognition ability make the new sensing system ideally suited for the measurement of BCAAs and the determination of the Fischer ratio, an indicator of hepatic disease involving metabolic dysfunction.
| S-EPMC3714611 | biostudies-literature
Project description:Four experiments were conducted to estimate the optimal standardized ileal digestible (SID) level of branched-chain amino acids in low-protein diets during the starter, grower, and finisher periods, using the response surface methodology, and to study their effects on performance and mRNA expression of genes involved in the mechanistic target of rapamycin (mTOR) pathway of broiler chickens from 8 to 21 D of age. In experiments 1, 2, and 3, a total of 1,500 Cobb male broiler chickens were assigned to 15 diets of a central composite rotatable design (CCD) of response surface methodology containing 5 levels of SID Leu, Val, and Ile with 5 replicate pens of 20 birds each. A 3-factor, 5-level CCD platform was used to fit the second-order polynomial equation of broiler performance. In experiment 4, a total of 540 8-day-old Cobb male broiler chickens were distributed in a completely randomized 2 x 3 x 3 factorial arrangement with 2 SID Leu levels (1.28 or 1.83%), 3 SID Val levels (0.65, 0.90, or 1.20%), and 3 SID Ile levels (0.54, 0.79, or 1.09%) for a total of 18 treatments with 5 replicate cages of 6 birds each. High Leu levels impaired (P < 0.05) gain:feed when birds were fed marginal Val or Ile diets. However, gain:feed was restored when both Val and Ile were supplemented to reach adequate or high levels. High Leu levels increased (P < 0.05) mRNA expression of S6K1 and eEF2 genes only in birds fed high Ile levels. Dietary SID Leu, Val, and Ile levels required for gain:feed optimization in low-protein diets were estimated at 1.37, 0.94, and 0.87% during the starter period; 1.23, 0.82, and 0.75% during the grower period; and 1.15, 0.77, and 0.70% during the finisher phase, respectively. Higher Val and Ile levels are required to optimize the effect of Leu supplementation on mRNA expression of mTOR pathway genes in the pectoralis major muscle of broilers from day 1 to 21 after hatch.
| S-EPMC7647919 | biostudies-literature
Project description:In this experiment we wanted to see how the binding behavior of the S. Cerevisiae transcription factor Leu3, on of the main regulators of leucine biosynthesis, is affected by different availability of the branched chain amino acids. For this we grow the cells in shake flask under glucose limitation and treated them 2 hours before sampling. The cells were then cross-linked with formaldehyde and ChIP-seq was performed using the Oxford Nanopore MinIon.
2020-07-01 | E-MTAB-8820 | biostudies-arrayexpress
Project description:BackgroundIn 2011-2012, Northern Vietnam experienced its first large scale hand foot and mouth disease (HFMD) epidemic. In 2011, a major HFMD epidemic was also reported in South Vietnam with fatal cases. This 2011-2012 outbreak was the first one to occur in North Vietnam providing grounds to study the etiology, origin and dynamic of the disease. We report here the analysis of the VP1 gene of strains isolated throughout North Vietnam during the 2011-2012 outbreak and before.MethodsThe VP1 gene of 106 EV-A71 isolates from North Vietnam and 2 from Central Vietnam were sequenced. Sequence alignments were analyzed at the nucleic acid and protein level. Gene polymorphism was also analyzed. A Factorial Correspondence Analysis was performed to correlate amino acid mutations with clinical parameters.ResultsThe sequences were distributed into four phylogenetic clusters. Three clusters corresponded to the subgenogroup C4 and the last one corresponded to the subgenogroup C5. Each cluster displayed different polymorphism characteristics. Proteins were highly conserved but three sites bearing only Isoleucine (I) or Valine (V) were characterized. The isoleucine/valine variability matched the clusters. Spatiotemporal analysis of the I/V variants showed that all variants which emerged in 2011 and then in 2012 were not the same but were all present in the region prior to the 2011-2012 outbreak. Some correlation was found between certain I/V variants and ethnicity and severity.ConclusionsThe 2011-2012 outbreak was not caused by an exogenous strain coming from South Vietnam or elsewhere but by strains already present and circulating at low level in North Vietnam. However, what triggered the outbreak remains unclear. A selective pressure is applied on I/V variants which matches the genetic clusters. I/V variants were shown on other viruses to correlate with pathogenicity. This should be investigated in EV-A71. I/V variants are an easy and efficient way to survey and identify circulating EV-A71 strains.
| S-EPMC5422960 | biostudies-literature
Project description:A methyl-detected 'out-and-back' NMR experiment for obtaining simultaneous correlations of methyl resonances of valine and isoleucine/leucine residues with backbone carbonyl chemical shifts, SIM-HMCM(CGCBCA)CO, is described. The developed pulse-scheme serves the purpose of convenience in recording a single data set for all Ile(δ1), Leu(δ) and Val(γ) (ILV) methyl positions instead of acquiring two separate spectra selective for valine or leucine/isoleucine residues. The SIM-HMCM(CGCBCA)CO experiment can be used for ILV methyl assignments in moderately sized protein systems (up to ~100 kDa) where the backbone chemical shifts of (13)C(α), (13)Cβ and (13)CO are known from prior NMR studies and where some losses in sensitivity can be tolerated for the sake of an overall reduction in NMR acquisition time.
| S-EPMC3974575 | biostudies-literature
Project description:A simple and cost effective method to independently and stereo-specifically incorporate [(1)H,(13)C]-methyls in Leu and Val in proteins is presented. Recombinant proteins for NMR studies are produced using a tailored set of auxotrophic E. coli strains. NMR active isotopes are routed to either Leu or Val methyl groups from the commercially available and scrambling-free precursors ?-ketoisovalerate and acetolactate. The engineered strains produce deuterated proteins with stereospecific [(1)H,(13)C]-methyl labeling separately at Leu or Val amino acids. This is the first method that achieves Leu-specific stereospecific [(1)H,(13)C]-methyl labeling of proteins and scramble-free Val-specific labeling. Use of auxotrophs drastically decreases the amount of labeled precursor required for expression without impacting the yield. The concept is extended to Thr methyl labeling by means of a Thr-specific auxotroph that provides enhanced efficiency for use with the costly L-[4-(13)C,2,3-(2)H2,(15)N]-Thr reagent. The Thr-specific strain allows for the production of Thr-[(13)CH3](?2) labeled protein with an optimal isotope incorporation using up to 50 % less labeled Thr than the traditional E. coli strain without the need for (2)H-glycine to prevent scrambling.
| S-EPMC4936824 | biostudies-literature
Project description:In maple syrup urine disease (MSUD), leucine (Leu) accumulation, and its metabolites cause brain toxicity, and at diagnosis rapid plasma Leu reduction is essential. Valine (Val) and isoleucine (Iso) supplements are necessary to promote anabolism and enable prompt reduction of plasma Leu. Val/Iso supplements are unavailable in Iran, so an alternative source was necessary. An emergency protocol was developed using an unconventional source of Val and Iso to prompt reduction of high plasma Leu levels during an acute metabolic crisis to prevent brain encephalopathy and neurological sequelae. Five children with classical MSUD were referred aged 1-25 months, with a prolonged high plasma Leu of more than 1500 μmol/L and acute symptoms (irritability, poor feeding, and hypotonia). Initially, breast milk/regular infant formula was stopped. Val and Iso were given in calculated amounts from a Leu-free formula containing Iso/Val (Xleu Maxamaid, Nutricia Ltd.) to promote anabolism. It was prescribed for a controlled and limited time with a branched chain amino acid (BCAA) free formula. Frequent amino acid monitoring was conducted. Natural protein was re-added after normalizing plasma Leu. Plasma Leu declined by a median (range) of 1677 (1501-1852) μmol/L within 3-4 days of intervention. The median follow-up time was 24 months (range: 14-32) and patients showed improvement in motor and cognitive skills after normalizing plasma Leu (75-200 μmol/L). Most had improvement in their head circumference (n = 4). Due to the unavailability of individual Val/Iso supplements, a Leu-free formula rapidly lowered plasma Leu concentrations during acute crisis, to prevent cerebral edema and brain damage in MSUD.
| S-EPMC9626667 | biostudies-literature
Project description:The reactions of [Ru(NO)Cl5](2-) with glycine (Gly), L-alanine (L-Ala), L-valine (L-Val), L-proline (L-Pro), D-proline (D-Pro), L-serine (L-Ser), L-threonine (L-Thr), and L-tyrosine (L-Tyr) in n-butanol or n-propanol afforded eight new complexes (1-8) of the general formula [RuCl3(AA-H)(NO)](-), where AA = Gly, L-Ala, L-Val, L-Pro, D-Pro, L-Ser, L-Thr, and L-Tyr, respectively. The compounds were characterized by elemental analysis, electrospray ionization mass spectrometry (ESI-MS), (1)H NMR, UV-visible and ATR IR spectroscopy, cyclic voltammetry, and X-ray crystallography. X-ray crystallography studies have revealed that in all cases the same isomer type (from three theoretically possible) was isolated, namely mer(Cl),trans(NO,O)-[RuCl3(AA-H)(NO)], as was also recently reported for osmium analogues with Gly, L-Pro, and D-Pro (see Z. Anorg. Allg. Chem. 2013, 639, 1590-1597). Compounds 1, 4, 5, and 8 were investigated by ESI-MS with regard to their stability in aqueous solution and reactivity toward sodium ascorbate. In addition, cell culture experiments in three human cancer cell lines, namely, A549 (nonsmall cell lung carcinoma), CH1 (ovarian carcinoma), and SW480 (colon carcinoma), were performed, and the results are discussed in conjunction with the lipophilicity of compounds.
| S-EPMC3942006 | biostudies-literature