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

Surveys of New Mexico Alfalfa Producers and Dairy Hay Users: Will Growth of the State's Dairy Industry be Limited by Alfalfa Availability?

Production Economics,

The Effect of Disclosure on Consumer Knowledge of Credit Terms: A Longitudinal Study

Early evaluations of Truth‐in‐Lending have observed impressive gains in consumer knowledge about interest rates. Contrary to original goals, consumers with more education, income, and debt experience have benefited far more than low‐income and minority consumers. How will these results change over time as consumers gain credit experience with the aid of disclosure? Has disclosure improved consumer understanding about finance charges, and what factors beyond socio‐economic status might have enhanced consumer knowledge of credit terms? These questions are addressed in this report of a large sample of California households surveyed at two points in time. The longitudinal analysis shows individual changes in knowledge, the effects of credit experience on learning, and a projection of future levels of credit knowledge

Mach-Zehnder Interferometry in a Strongly Driven Superconducting Qubit

We demonstrate Mach-Zehnder-type interferometry in a superconducting flux qubit. The qubit is a tunable artificial atom, whose ground and excited states exhibit an avoided crossing. Strongly driving the qubit with harmonic excitation sweeps it through the avoided crossing two times per period. As the induced Landau-Zener transitions act as coherent beamsplitters, the accumulated phase between transitions, which varies with microwave amplitude, results in quantum interference fringes for n=1...20 photon transitions. The generalization of optical Mach-Zehnder interferometry, performed in qubit phase space, provides an alternative means to manipulate and characterize the qubit in the strongly-driven regime.Comment: 14 pages, 6 figure

Ohio Livestock Waste Management Guide

PDF pages: 3

The Effects of Supplementing Wet Distillers Grains Mixed With Wheat Straw to Growing Steers

A growing study compared feeding wet distillers grains, dried distillers grains, and a mix of 66% wet distillers grains and 33% wheat straw as supplements to a forage-based diet. Steers were supplemented 0, 2, 4, or 6 lb distillers grains/head daily. Wet distillers grains and dried distillers grains produced higher final body weight and dry matter intake compared to the mix. Increasing levels of distillers grains increased performance in forage based diets and wet grains mixed with straw reduced forage intake

A Computational Model of Driver Decision Making at an Intersection Controlled by a Traffic Light

An important challenge associated with driving simulation development is the computational representation of agent behaviors. This paper describes the development of a preliminary autonomous agent behavior model (based on the Recognition-Primed Decision (RPD) model, and Hintzman’s multiple-trace memory model) mimicking human decision making in approaching an intersection controlled by a traffic light. To populate the model, an initial Cognitive Task Analysis was conducted with six drivers to learn the important cues, expectancies, goals, and courses of action associated with traffic light approach. The agent model learns to associate environmental cues (such as traffic light color) with expectancies of upcoming events (like light color change) and appropriate courses of action (such as decelerating). At present, the model is currently being evaluated for its successful representation of the RecognitionPrimed Decision Making process

Resonant Readout of a Persistent Current Qubit

We have implemented a resonant circuit that uses a SQUID as a flux-sensitive Josephson inductor for qubit readout. In contrast to the conventional switching current measurement that generates undesired quasi-particles when the SQUID switches to the voltage state, our approach keeps the readout SQUID biased along the supercurrent branch during the measurement. By incorporating the SQUID inductor in a high-Q resonant circuit, we can distinguish the two flux states of a niobium persistent-current (PC) qubit by observing a shift in the resonant frequency of both the magnitude and the phase spectra. The readout circuit was also characterized in the nonlinear regime to investigate its potential use as a nonlinear amplifier.Comment: 4 pages, 2004 ASC Proceeding

The Effects of Supplementing Wet Distillers Grains Mixed With Wheat Straw to Growing Steers

A growing study compared feeding wet distillers grains, dried distillers grains, and a mix of 66% wet distillers grains and 33% wheat straw as supplements to a forage-based diet. Steers were supplemented 0, 2, 4, or 6 lb distillers grains/head daily. Wet distillers grains and dried distillers grains produced higher final body weight and dry matter intake compared to the mix. Increasing levels of distillers grains increased performance in forage based diets and wet grains mixed with straw reduced forage intake

Characteristics of Manure Harvested from Beef Cattle Feedlots

Open lot cattle production systems present unique challenges for nutrient planning processes. Previous estimates of quantities and characteristics of harvested manure from this type of facility are based upon data from the early 1970s. In addition, harvested manure is impacted by weather, feeding program, season, and pen management decisions. The objectives of this study are to characterize under commercial conditions for open lot beef systems: 1) harvested manure quantities and characteristics; 2) impact of factors such as feeding program, season, and management on harvested manure; and 3) mass balance for nitrogen (N) and phosphorus (P). Data from six commercial feedlots (representing 6,366 head of cattle) suggest that 33% of excreted N (65 g/hd/d) and 91% of excreted P (32 g/hd/d) are harvested as manure on average and that current standard estimates published by ASAE (2005) and NRCS (1992a) overestimate harvested manure N and P. Additionally, significant variation was observed among feedlots. This variation is driven by ration nutrient concentration (P only), pen conditions prior to and during manure harvest (N and P), and management choices relative to use of manure in lot maintenance (N and P). The variation would suggest that nutrient planning estimates for open lots would need to be based upon farm specific data as opposed to typical or standard values. Finally, a pen-based mass nutrient balance for a beef cattle feedlot suggests that pen outputs as finished animal, harvested manure, and nutrient losses represent 31%, 23%, and 47%, respectively, of all pen N inputs and 38%, 57%, and 5%, respectively, of all pen P inputs. Inputs include nutrient content of all animals and feed entering a feedlot pen over a grow-out period