Structure, substrate recognition and reactivity of Leishmania major mevalonate kinase

This research was supported by the German Academic Exchange Service (DAAD), the Wellcome Trust (TKS and WNH as Trust Senior Research fellows), the Biotechnology and Biological Science Research Council (Structural Proteomics of Rational Targets) and the European Synchrotron Radiation Facility.Background: Isoprenoid precursor synthesis via the mevalonate route in humans and pathogenic trypanosomatids is an important metabolic pathway. There is however, only limited information available on the structure and reactivity of the component enzymes in trypanosomatids. Since isoprenoid biosynthesis is essential for trypanosomatid viability and may provide new targets for therapeutic intervention it is important to characterize the pathway components. Results: Putative mevalonate kinase encoding genes from Leishmania major (LmMK) and Trypanosoma brucei (TbMK) have been cloned, over-expressed in and proteins isolated from procyclic-form T. brucei. A highly sensitive radioactive assay was developed and shows ATP-dependent phosphorylation of mevalonate. Apo and (R)-mevalonate bound crystal structures of LmMK, from a bacterial expression system, have been determined to high resolution providing, for the first time, information concerning binding of mevalonate to an MK. The mevalonate binds in a deep cavity lined by highly conserved residues. His25 is key for binding and for discrimination of (R)-over (S)-mevalonate, with the main chain amide interacting with the C3 hydroxyl group of ( R)mevalonate, and the side chain contributing, together with Val202 and Thr283, to the construction of a hydrophobic binding site for the C3 methyl substituent. The C5 hydroxyl, where phosphorylation occurs, points towards catalytic residues, Lys18 and Asp155. The activity of LmMK was significantly reduced compared to MK from other species and we were unable to obtain ATP-binding data. Comparisons with the rat MK:ATP complex were used to investigate how this substrate might bind. In LmMK, helix alpha 2 and the preceding polypeptide adopt a conformation, not seen in related kinase structures, impeding access to the nucleotide triphosphate binding site suggesting that a conformational rearrangement is required to allow ATP binding. Conclusion: Our new structural information, consistent with data on homologous enzymes allows a detailed description of how mevalonate is recognized and positioned for catalysis in MK. The mevalonate-binding site is highly conserved yet the ATP-binding site is structurally distinct in LmMK. We are unable to provide a definitive explanation for the low activity of recombinant protein isolated from a bacterial expression system compared to material isolated from procyclic-form Trypanosoma brucei.Publisher PDFPeer reviewe

A memodyne 3722 cassette reader system for the HP 2100 computer and program carp

This report describes a special purpose electronic interface between the Mernodyne 3722 High Speed cassette Tape Reader and the Hewlett Packard 2100 series computer with program CARP using an HP 12566 interface. A description of the operation and alignment procedures is included.Prepared for the Office of Naval Research under Contract N00014-76-C-0197; NR 083-004

Freedom and Excellence

Commencement address given by Terry Wickham, President of Heidelberg College, to the Winter 1959 graduating class of The Ohio State University, St. John Arena, Columbus, Ohio, March 19, 1959

Theoretical interpretation of scanning tunneling microscopy images: Application to the molybdenum disulfide family of transition metal dichalcogenides

We have performed ab initio quantum mechanical calculations to describe scanning tunneling microscopy (STM) images of MoS_2 and MoTe_2. These results indicate that the interpretation of the STM images of these and related materials depends sensitively on experimental conditions. For example, determining whether the maximum tunneling current correlates to the top atom (S or Te) or to the second‐layer atom (Mo) requires information on the tip‐sample separation. Based on these results we discuss some STM experimental procedures which would allow assignment of the chemical identity of STM spots with greater certainty

The Conductance of Some Non-Rigid Bolaform Electrolytes

Bolaform electrolytes are those which contain two or more charges separated by an inert framework. These are intermediate in structure between simple electrolytes and polyelectrolytes. Since these are similar to both synthetic and biological electrolytes, this investigation has been undertaken to determine some of their properties in solution. The sodium and magnesium salts of ethane, propane, butane, pentane, hexane and decane α,ω-disulfonates were prepared. Each acid was prepared by refluxing the α,ω-dibromide of the desired salt with sodium sulfite. The sodium and magnesium salts were prepared by ion-exchange techniques. The conductance of each salt was determined in methanol utilizing the weight dilution method. A data treatment method for unsymmetrical 1-2 electrolytes was tested. The Murphy-Cohen equation: ∧ = ∧° - S√C + E′ClnC + J′C was found to be invalid in the concentration range studied, 1.0 - 5.0 x 10-4moles/liter. The Onsager limiting law when applied to the sodium salt data gave ∧° values from approximately 108 (ohm-1cm2equiv-1) for the ethane salt to 100 (ohm-1cm2equiv-1) for the decane. Phoreograms showed these salts to be unassociated in methanol. The magnesium salts deviated substantially below the limiting law plot and consequently were considered highly associated. The ∧° values for the magnesium salts were calculated from Kohlrausch\u27s law, ∧° = λ+Mg + λ-anion. By fixing this value, association constants were found to be in a range of 108,950 for the ethane salt to 25,260 for the decane salt. These values show a monotonic decrease as the chain length of the salt increases. This is explained by charge separation considerations. As the chain length increases the charge density on the molecule decreases, illustrating a decrease in ion-ion interactions. The frictional coefficients of the sodium salts were determined from λ- data. These showed values of .0496 X 10-7 g/sec for the ethane salt to .0563 X 10-7 g/sec for decane salts. The Peterlin rod and bead model for the determination of frictional forces was successfully applied to the data. The Perrin rigid ellipsoid model did not show any agreement when experimental and theoretical frictional coefficient values were compared

Evaluation of a Ninth Grade Transition Program for At-Risk Students.

Researchers have identified the transition from middle to high school as a critical point in teenagers\u27 educational development. Despite sweeping educational reforms, many students are leaving high school without graduating. Local school districts are struggling to redesign high schools to provide the educational programs necessary for at-risk students\u27 success. Creating smaller learning communities and developing transition programs have shown success in addressing these problems. Sustained research shows promise in minimizing the effects of the transition process and adds to the body of knowledge. A case study approach using archival data was used to explore the differences in three Bearcat PRIDE treatment groups and their 8th (pre-treatment) and 9th (post-treatment) grade measures. Results of this study suggested students who participated in the Bearcat PRIDE program showed greater success in academic performance, reduced disciplinary actions, increased school attendance, and participation in extra-curricular activities of their 8th (pre-treatment) and 9th (post-treatment) grade measures

Iron vs. gold : a study of the three Anglo-Dutch wars, 1652-1674

The purpose of this paper is to show that, as the result of twenty-two years of intermittent warfare between England and the Netherlands, the English navy became established as the primary naval power of Europe. Also, I intend to illustrate that, as a by-product of this naval warfare, Dutch trade was seriously hurt, with the·major benefactors of this Dutch loss of trade being the English. This paper grew out of a seminar paper on the first Anglo-Dutch war for a Tudor and Stuart English History graduate seminar class taught in the fall of 1966 by Dr. John R. Rilling of the University or Richmond. Because in the present paper I attempt to cover such a large topic, all other aspects of English history of this period will be covered only insofar as they affected the Dutch wars