Synthesis Of N-Labeled Peptidyl AMP

Abstract

This paper deals with the synthesis of a new type of N-labeled peptidyl AMP, which would be used as a good substrate for analysis of the peptidyl transfer reaction on ribosome and for co-crystallization with ribosome. 4-(Dimethylamino)azobenzene-4′-sulfonyl (Dabsyl) was selected as the labeling group. (N-Dabsylglycyl)-L-leucyl AMP was synthesized from glycyl-L-leucine via a three-step procedure.     

Molecular cloning of cDNAs and genes for three alpha-glucosidases from European honeybees, Apis mellifera L., and heterologous production of recombinant enzymes in Pichia pastoris

cDNAs encoding three alpha-glucosidases (HBGases I, II, and III) from European honeybees, Apis mellifera, were cloned and sequenced, two of which were expressed in Pichia pastoris. The cDNAs for HBGases I, II, and III were 1,986, 1,910, and 1,915 bp in length, and included ORFs of 1,767, 1,743, and 1,704 bp encoding polypeptides comprised of 588, 580, and 567 amino acid residues, respectively. The deduced proteins of HBGases I, II, and III contained 18, 14, and 8 putative N-linked glycosylation sites, respectively, but at least 2 sites in HBGase II were unmodified by N-linked oligosaccharide. In spite of remarkable differences in the substrate specificities of the three HBGases, high homologies (38-44% identity) were found in the deduced amino acid sequences. In addition, three genomic DNAs, of 13,325, 2,759, and 27,643 bp, encoding HBGases I, II, and III, respectively, were isolated from honeybees, and the sequences were analyzed. The gene of HBGase I was found to be composed of 8 exons and 7 introns. The gene of HBGase II was not divided by intron. The gene of HBGase III was confirmed to be made up of 9 exons and 8 introns, and to be located in the region upstream the gene of HBGase I.     

The difference in output properties between dominant and nondominant limbs as measured by various muscle function tests

Recently, right and left output properties exerted from specific muscle groups have been evaluated using special measurement devices such as an isokinetic dynamometer. However, it remains unclear whether the coach can properly evaluate muscle function corresponding to lateral specificity in athletes. This study aimed to examine the different output properties between the dominant (D) and nondominant (ND) upper limbs as measured by muscle function tests with various muscle contraction patterns. Fifteen right-handed young men participated in this study. Each subject carried out isometric, isokinetic, and isotonic muscle power tests by elbow flexion with right and left arms. When calculating the laterality index, the laterality of the isotonic test (1.17) was the highest. In all tests, significant correlations were found between the measurements of the D and ND limbs. The isometric test was the highest (r = 0.93), followed by the isokinetic test (r = 0.66-0.83) and the isotonic test (r = 0.55). To examine the ratio of the laterality of measurements provided by each muscle function test, each measurement was converted to a standard score (Z-score). There were significant differences between D and ND limbs in the isometric (D:ND = 55.0:45.0) and the isotonic (54.1:45.9) tests but not in the isokinetic test (60°·s?1, 51.4:48.6; 180°·s?1, 50.7:49.2; 300°·s?1, 51.8:48.2). Particularly, the D (right) limb exerted greater muscle power in the isometric and the isotonic tests than in the isokinetic test. Occupational therapists or strength and conditioning professionals should understand that the D-ND differences shown by these muscle function tests may differ because of measurement conditions.     

High-performance liquid chromatography followed by peroxyoxalate chemiluminescence detection of acetylcholine and choline utilizing immobilized enzymes

A sensitive and selective method for the simultaneous determination of acetylcholine (ACh) and choline (Ch) is reported. ACh and Ch were separated on a reversed-phase column, passed through an immobilized enzymes (acetylcholine esterase and choline oxidase) column, and converted to hydrogen peroxide. The generated hydrogen peroxide was detected by the peroxyoxalate chemiluminescence reaction. The linear determination ranges were from 10 pmol to 10 nmol. The detection limit for both cholines was 1 pmol.     

Control scheme of two d.o.f. CPM device to suppress the extension of ligament of the elbow

Continuous passive motion (CPM) is an orthopedic treatment or a physiotherapy and has been applied after surgery. After surgery for the injured ulna collateral ligament (UCL) which is a major clinical case in the elbow joint, the reaction force at the hand of the patient increases and may be excessively large due to an increase of the joint stiffness. For this, it will be effective to reduce the excessive reaction force by controlling pro-/supination of forearm. The UCL in the elbow joint is extended due to pro-/supination of forearm, but the excessive extension may aggravate the injury of the UCL. Thus, it is required to suppress both the excessive extension of the UCL and the reaction force. In this paper, a control scheme of CPM device that can suppress both the excessive extension of the UCL and the reaction force is proposed, and its effectiveness is confirmed from the experimental results.     

Metabolomic analyses of plasma and liver of mice fed with immature Citrus tumida peel

ABSTRACT

In this study, we investigated the effects of dietary supplementation of Citrus tumida hort. ex Tanaka on food intake, body and fat tissue weights, and metabolic profiles of plasma and liver in mice. Supplementation with 5% (w/w) of peels of immature C. tumida (PIC) for 4 weeks significantly suppressed body weight gain and decreased adipose tissue weight in epididymal, perirenal, and subcutaneous fats. Metabolome analyses showed that 2-hydroxyvaleric acid levels were reduced in the blood plasma of mice fed with PIC. PIC supplementation significantly elevated dipeptide (Thr-Asp, Ser-Glu, and Ala-Ala), glucuronic acid, and S-methylglutathione levels, and significantly reduced betaine aldehyde levels in the liver. In conclusion, PIC supplementation affects the metabolism of fatty acids, pectin, glutathione, and choline, showing potential beneficial effects for metabolic syndrome and obesity. PIC may be developed as a functional food and used in the treatment of these diseases.     

Improved Polymerase Chain Reaction-restriction Fragment Length Polymorphism Genotyping of Toxic Pufferfish by Liquid Chromatography/Mass Spectrometry

An improved version of a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method for genotyping toxic pufferfish species by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) is described. DNA extraction is carried out using a silica membrane-based DNA extraction kit. After the PCR amplification using a detergent-free PCR buffer, restriction enzymes are added to the solution without purifying the reaction solution. A reverse-phase silica monolith column and a Fourier transform high resolution mass spectrometer having a modified Kingdon trap analyzer are employed for separation and detection, respectively. The mobile phase, consisting of 400 mM 1,1,1,3,3,3-hexafluoro-2-propanol, 15 mM triethylamine (pH 7.9) and methanol, is delivered at a flow rate of 0.4 ml/min. The cycle time for LC/ESI-MS analysis is 8 min including equilibration of the column. Deconvolution software having an isotope distribution model of the oligonucleotide is used to calculate the corresponding monoisotopic mass from the mass spectrum. For analysis of oligonucleotides (range 26-79 nucleotides), mass accuracy was 0.62 ± 0.74 ppm (n = 280) and excellent accuracy and precision were sustained for 180 hr without use of a lock mass standard.