Reductive Alkylation of Proteins Towards Structural and Biological Applications

Nuclear magnetic resonance (NMR) spectroscopy is a proven technique for protein structure and dynamic studies, typically requiring the incorporation of stable magnetic isotopes to improve sensitivity and assign resonances. Degenerate levels of 13C-incorporation have been the biggest obstacle for mass spectrometry-assisted assignment of 13C-dimethylamine resonances in nuclear magnetic resonance spectroscopy (NMR). Reductive 13C-methylation is an alternative labeling method for proteins not amenable to bacterial host overexpression. Because reductive 13C-methylation adds sparse, isotopic labels, traditional methods of assigning the NMR signals are not applicable. The research presented in the first part of this dissertation explores several methods used to break the degeneracy in 13C-labeling of lysozyme. To overcome the degeneracy in labeling lysozyme with the reductive methylation reaction, we investigated two methods: 1) reductive methylation in the presence of 18-crown-6-ether (18C6) and 2) reductive methylation using multiple reducing agents. To assign the - and -dimethylamine resonances of the N-terminal lysine residue of lysozyme, a non-destructive Edman degradation method was explored. The second part of this research discusses an alternative assignment method based on mass spectrometry to aid in the assignment of the NMR signals from reductively 13C-methylated proteins. Because assignment is increasingly difficult when lysine is the N-terminal residue of the protein, one method is described to identify the NMR resonance of the 13C-methyls associated with both the N-terminal α-amine and the side chain ε-amine. The NMR signals of the N-terminal α-dimethylamine and the side chain ε-dimethylamine of hen egg white lysozyme Lys1 are identified in 1H-13C heteronuclear single-quantum correlation spectra. Protein chemical modification is a well-established field that continues to impact leading research today including glycomimetics and cross-linking of proteins. Current protein chemical modifications like polyethylene glycol are proven useful for increasing the lifetime of several therapeutic enzymes but are also toxic to the body. In the last chapter, we present the use of sugar derivatives as a possible less toxic alternative for synthetic glycoproteins. The synthesis of a protein modifier is described and preliminary data of its application as a glycomimetic and cross-linking agent is presented

Trustworthy Experimentation Under Telemetry Loss

Failure to accurately measure the outcomes of an experiment can lead to bias and incorrect conclusions. Online controlled experiments (aka AB tests) are increasingly being used to make decisions to improve websites as well as mobile and desktop applications. We argue that loss of telemetry data (during upload or post-processing) can skew the results of experiments, leading to loss of statistical power and inaccurate or erroneous conclusions. By systematically investigating the causes of telemetry loss, we argue that it is not practical to entirely eliminate it. Consequently, experimentation systems need to be robust to its effects. Furthermore, we note that it is nontrivial to measure the absolute level of telemetry loss in an experimentation system. In this paper, we take a top-down approach towards solving this problem. We motivate the impact of loss qualitatively using experiments in real applications deployed at scale, and formalize the problem by presenting a theoretical breakdown of the bias introduced by loss. Based on this foundation, we present a general framework for quantitatively evaluating the impact of telemetry loss, and present two solutions to measure the absolute levels of loss. This framework is used by well-known applications at Microsoft, with millions of users and billions of sessions. These general principles can be adopted by any application to improve the overall trustworthiness of experimentation and data-driven decision making.Comment: Proceedings of the 27th ACM International Conference on Information and Knowledge Management, October 201

Analysis of operational risk of banks – catastrophe modelling

Nowadays financial institutions due to regulation and internal motivations care more intensively on their risks. Besides previously dominating market and credit risk new trend is to handle operational risk systematically. Operational risk is the risk of loss resulting from inadequate or failed internal processes, people and systems or from external events. First we show the basic features of operational risk and its modelling and regulatory approaches, and after we will analyse operational risk in an own developed simulation model framework. Our approach is based on the analysis of latent risk process instead of manifest risk process, which widely popular in risk literature. In our model the latent risk process is a stochastic risk process, so called Ornstein- Uhlenbeck process, which is a mean reversion process. In the model framework we define catastrophe as breach of a critical barrier by the process. We analyse the distributions of catastrophe frequency, severity and first time to hit, not only for single process, but for dual process as well. Based on our first results we could not falsify the Poisson feature of frequency, and long tail feature of severity. Distribution of “first time to hit” requires more sophisticated analysis. At the end of paper we examine advantages of simulation based forecasting, and finally we concluding with the possible, further research directions to be done in the future

Controlling evaporation loss from water storages

[Executive Summary]: Evaporation losses from on-farm storage can potentially be large, particularly in irrigation areas in northern New South Wales and Queensland where up to 40% of storage volume can be lost each year to evaporation. Reducing evaporation from a water storage would allow additional crop production, water trading or water for the environment. While theoretical research into evaporation from storages has previously been undertaken there has been little evaluation of current evaporation mitigation technologies (EMTs) on commercial sized water storages. This project was initiated by the Queensland Government Department of Natural Resources and Mines (NRM) with the express aim of addressing this gap in our knowledge. The report addressed i) assessment of the effectiveness of different EMT’s in reducing evaporation from commercial storages across a range of climate regions, ii) assessment of the practical and technical limitations of different evaporation control products, and iii) comparison of the economics of different EMT’s on water storages used for irrigation

Hypermedia-based discovery for source selection using low-cost linked data interfaces

Evaluating federated Linked Data queries requires consulting multiple sources on the Web. Before a client can execute queries, it must discover data sources, and determine which ones are relevant. Federated query execution research focuses on the actual execution, while data source discovery is often marginally discussed-even though it has a strong impact on selecting sources that contribute to the query results. Therefore, the authors introduce a discovery approach for Linked Data interfaces based on hypermedia links and controls, and apply it to federated query execution with Triple Pattern Fragments. In addition, the authors identify quantitative metrics to evaluate this discovery approach. This article describes generic evaluation measures and results for their concrete approach. With low-cost data summaries as seed, interfaces to eight large real-world datasets can discover each other within 7 minutes. Hypermedia-based client-side querying shows a promising gain of up to 50% in execution time, but demands algorithms that visit a higher number of interfaces to improve result completeness

Empirical study on intrinsic motivation factors of employees in transition economies

Purpose: This research study aims to unequivocally discuss and explore the intrinsic motivation factors of the employees in transition economies. The purpose of this study is to find which are the main internal factors that motivate employees? Taking in to consideration changes to the economic systems, different factors derived from theories, as well as other research analyzed. Design/Methodology/Approach: To describe the relationship between motivational factors that motivate workers today, we have developed a questionnaire which is distributed and filled by 510 respondents, not taking into account their hierarchy in the organization. Methodologies used in this research are Pearson correlation matrix to test the relationship with independent variables and dependent variable which in our case is intrinsic motivation and as a main econometric model binary logit is used Findings: The research shows that most of the data that we have used in this model are correlated, and have high level of significance, and there is no multicollinearity. Whereas, in Binary Logit model used Cox & Snell R Square is .696 meaning that our model is predicted by around 70 percent from the independent variables. From our research we found that most important intrinsic motivational factors are competence, job satisfaction, job recognition as well as financial rewards. There is no impact on intrinsic motivation that is dependent on finding the job interesting or not. Worthy of note is that the odds of employees to be intrinsically motivated are much higher in public institution than in private businesses. Practical Implications: This study raises awareness of motivational factors in economies that are still in transition. Results of this study can serve as a good guide especially for small businesses and business startups on importance of behaviorist approach for motivation factors as competence and job recognition. Originality/Value: this paper provides original insights of the compound relationship between different factors that have a direct affect in intrinsic motivation of employees’ therefore as an outcome better job performance.peer-reviewe