Differences in folate-protein interactions result in differing inhibition of native rat liver and recombinant glycine N-methyltransferase by 5-methyltetrahydrofolate

Glycine N-methyltransferase (GNMT) is a key regulatory enzyme in methyl group metabolism. In mammalian liver it reduces S-adenosylmethionine levels by using it to methylate glycine, producing N-methylglycine (sarcosine) and S-adenosylhomocysteine. GNMT is inhibited by binding two molecules of 5-methyltetrahydrofolate (mono- or polyglutamate forms) per tetramer of the active enzyme. Inhibition is sensitive to the status of the N-terminal valine of GNMT and to polyglutamation of the folate inhibitor. It is inhibited by pentaglutamate form more efficiently compared to monoglutamate form. The native rat liver GNMT contains an acetylated N-terminal valine and is inhibited much more efficiently compared to the recombinant protein expressed in E. coli where the N-terminus is not acetylated. In this work we used a protein crystallography approach to evaluate the structural basis for these differences. We show that in the folate-GNMT complexes with the native enzyme, two folate molecules establish three and four hydrogen bonds with the protein. In the folate-recombinant GNMT complex only one hydrogen bond is established. This difference results in more effective inhibition by folate of the native liver GNMT activity compared to the recombinant enzyme. © 2011 Elsevier B.V. All rights reserved

An Inquiry into the Aviation Management Education Paradigm Shift

Working adults with four-year degrees from accredited colleges or universities earn, on average, almost three times more than individuals without a degree. This pay gap led Newcomer and his colleagues to study attitudes of aviation and aerospace managers towards education. That study found that managers valued education in new hires, even though they did not deem it critical to their own positions. That finding indicated a potential paradigm shift towards the perceived value of education in the industry. In the current qualitative, phenomenological research, we interviewed 14 managers from various capacities within the aviation and aerospace industries to determine the relative importance of education, certification, and experience when hiring or selecting new team members. The results indicated that managers value experience most when making staffing decisions. Next, they value certification or education, depending on the technical or managerial role. A majority of the managers did express that their attitudes towards the value of education had become stronger over their careers. The study has hiring implications for aviation and aerospace managers, as well as employees, in terms of what to focus on in interviews and in reviewing candidate credentials

Folate in demethylation: The crystal structure of the rat dimethylglycine dehydrogenase complexed with tetrahydrofolate

Dimethylglycine dehydrogenase (DMGDH) is a mammalian mitochondrial enzyme which plays an important role in the utilization of methyl groups derived from choline. DMGDH is a flavin containing enzyme which catalyzes the oxidative demethylation of dimethylglycine in vitro with the formation of sarcosine (N-methylglycine), hydrogen peroxide and formaldehyde. DMGDH binds tetrahydrofolate (THF) in vivo, which serves as an acceptor of formaldehyde and in the cell the product of the reaction is 5,10-methylenetetrahydrofolate instead of formaldehyde. To gain insight into the mechanism of the reaction we solved the crystal structures of the recombinant mature and precursor forms of rat DMGDH and DMGDH-THF complexes. Both forms of DMGDH reveal similar kinetic parameters and have the same tertiary structure fold with two domains formed by N- and C-terminal halves of the protein. The active center is located in the N-terminal domain while the THF binding site is located in the C-terminal domain about 40 Å from the isoalloxazine ring of FAD. The folate binding site is connected with the enzyme active center via an intramolecular channel. This suggests the possible transfer of the intermediate imine of dimethylglycine from the active center to the bound THF where they could react producing a 5,10- methylenetetrahydrofolate. Based on the homology of the rat and human DMGDH the structural basis for the mechanism of inactivation of the human DMGDH by naturally occurring His109Arg mutation is proposed. ©2014 Elsevier Inc. All rights reserved

5-Methyltetrahydrofolate is bound in intersubunit areas of rat liver folate-binding protein glycine N-methyltransferase

Glycine N-methyltransferase (GNMT) is a key regulatory enzyme in methyl group metabolism. It is abundant in the liver, where it uses excess S-adenosylmethionine (AdoMet) to methylate glycine to N-methylglycine (sarcosine) and produces S-adenosylhomocysteine (AdoHcy), thereby controlling the methylating potential of the cell. GNMT also links utilization of preformed methyl groups, in the form of methionine, to their de novo synthesis, because it is inhibited by a specific form of folate, 5-methyltetrahydrofolate. Although the structure of the enzyme has been elucidated by x-ray crystallography of the apoenzyme and in the presence of the substrate, the location of the folate inhibitor in the tetrameric structure has not been identified. We report here for the first time the crystal structure of rat GNMT complexed with 5-methyltetrahydrofolate. In the GNMT-folate complex, two folate binding sites were located in the intersubunit areas of the tetramer. Each folate binding site is formed primarily by two 1-7 N-terminal regions of one pair of subunits and two 205-218 regions of the other pair of subunits. Both the pteridine and p-aminobenzoyl rings are located in the hydrophobic cavities formed by Tyr 5, Leu207, and Met215 residues of all subunits. Binding experiments in solution also confirm that one GNMT tetramer binds two folate molecules. For the enzymatic reaction to take place, the N-terminal fragments of GNMTmust have a significant degree of conformational freedom to provide access to the active sites. The presence of the folate in this position provides a mechanism for its inhibition

Validating the Knowledge, Skills, and Abilities Composite Measure: An Aviation Industry Pilot Study

Recent research into the aviation management education paradigm shift indicated that education, certification, and experience (ECE) were all important in the aviation industry; however, the relative importance varied between managers depending on their field. Overall, the results identified experience as the most important factor, followed by certification, then education. Furthermore, the authors made a logical connection between the that study and knowledge, skills, and abilities (KSAs), which would ultimately lead managers to a hiring decision. The purpose of this mixed-methods sequential exploratory pilot study was to develop, validate, and test the reliability of the KSA composite measure (KCM), a data collection device to measure the connection between ECE and KSAs. The three phased approach consisted of (a) an expert panel review by eight research, aviation, and human resource professionals, (b) an institutional review board (IRB) review, and (c) a live pilot using 45 professionals from the aviation industry. The pilot proved to validate and confirm the reliability of the KCM for future use

Hydrogeologic controls on ground-water and contaminant discharge to the Columbia River near the Hanford Townsite

The purpose of this study is to quantify ground-water and contaminant discharge to the Columbia River in the Hanford Townsite vicinity. The primary objectives of the work are to: describe the hydrogeologic setting and controls on ground-water movement and contaminant discharge to the Columbia River; understand the river/aquifer relationship and its effects on contaminant discharge to the Columbia River; quantify the ground-water and contaminant mass discharge to the Columbia River; and provide data that may be useful for a three-dimensional model of ground-water flow and contaminant transport in the Hanford Townsite study area. The majority of ground-water contamination occurs within the unconfined aquifer; therefore, ground-water and contaminant discharge from the unconfined aquifer is the emphasis of this study. The period of study is primarily from June 1990 through March 1992

The HADES Tracking System

The tracking system of the dielectron spectrometer HADES at GSI Darmstadt is formed out of 24 low-mass, trapezoidal multi-layer drift chambers providing in total about 30 square meter of active area. Low multiple scattering in the in total four planes of drift chambers before and after the magnetic field is ensured by using helium-based gas mixtures and aluminum cathode and field wires. First in-beam performance results are contrasted with expectations from simulations. Emphasis is placed on the energy loss information, exploring its relevance regarding track recognition.Comment: 6 pages, 4 figures, presented at the 10th Vienna Conference on Instrumentation, Vienna, February 2004, to be published in NIM A (special issue

A double-sided, shield-less stave prototype for the ATLAS upgrade strip tracker for the high luminosity LHC

A detailed description of the integration structures for the barrel region of the silicon strips tracker of the ATLAS Phase-II upgrade for the upgrade of the Large Hadron Collider, the so-called High Luminosity LHC (HL-LHC), is presented. This paper focuses on one of the latest demonstrator prototypes recently assembled, with numerous unique features. It consists of a shortened, shield-less, and double sided stave, with two candidate power distributions implemented. Thermal and electrical performances of the prototype are presented, as well as a description of the assembly procedures and tools