The Modification and Role of a Methyl-Arginine Post-Translational Modification in the Active Site of the Enzyme Methyl-Coenzyme M Reductase

The enzyme methyl-coenzyme M reductase (Mcr) is used in methanogenic archaea to produce methane and in anaerobic methanotrophs to oxidize methane. A significant portion of the methane produced by methanogens is released to the atmosphere, where it contributes to global warming. Understanding Mcr could lead to the inhibition of methane production or the optimization of the enzyme to start the conversion of methane into biofuels. Chapter 1 provides a background to the work that will be discussed in this dissertation. Chapter 2 examines methanogenesis marker protein 10 (Mmp10), a proposed radical SAM enzyme that may modify an arginine residue within Mcr. Multiple in vivo studies have examined how knocking out Mmp10 affects whole cells. Here, in vitro experiments with Mmp10 are used to study its methylating ability and investigate the cofactors present in this enzyme. Mass spectrometry show that Mmp10 can methylate the target arginine within a substrate peptide modeled after Mcr. Spectroscopy reveals that Mmp10 contains both a [4Fe-4S](II)/(I) cluster and a cobamide cofactor. In Chapter 3, the activation of Mcr in Methanococcus maripaludis is studied. Mcr is only active in the Ni(I) (Mcr-red1) form, but purifying Mcr without pretreatment produces only the Ni(II) (Mcr-silent) form. Purified Mcrsilent cannot be converted chemically to Mcrred1; however, the Ni(III) (Mcr-ox1) form of Mcr can be reduced to Mcr-red1 in vitro. These experiments examine how the Mcr-ox1 or Mcr-red1 forms can be induced in the whole cells through incubation with sodium sulfide or 100% H2, respectively. As shown by spectroscopy, Mcr-red1 can be induced in whole cells grown on 80% H2/20% CO2 and incubated with 100% H2; however, Mcrox1 cannot be induced when formate-grown cells are incubated with sodium sulfide. Chapter 4 investigates the iron-sulfur cluster content in a radical SAM enzyme, MptM. The primary sequence of MptM reveals two CX3CX2C motifs that could bind SAM clusters and other potential binding sites for [4Fe-4S](II)/(I) clusters. Site-directed mutagenesis was performed on multiple potential cluster sites and the samples were analyzed with electron paramagnetic resonance (EPR) spectroscopy. The data reveals that there are three [4Fe-4S](II)/(I) clusters within MptM

Three-dimensional numerical simulations of subaerial landslide generated waves

This research aims to advance the continuing effort of general purpose computational fluid dynamics model validation of subaerial landslide generated wave (SLGW) simulations. Specifically, using the open source program weakly compressible Smooth Particle Hydrodynamics model, DualSPHysics, three-dimensional simulations are quantitatively compared against a combination of physical model data and traditional general-purpose computational fluid dynamics, Flow-3D™, data. Many simulations were conducted to determine the effect of both numerical parametrization and numerical scheme prescriptions on SLGW accuracy. A systematic approach was taken to parse out insignificant physical processes using Flow-3D™ - specifically surface tension - and to determine the optimal numerical scheme settings that yield the most accurate results for both Flow-3D™ and DualSPHysics. From this research, it is found that DualSPHysics is able to accurately simulate both wave generation and wave propagation, but tends to over-predict the maximum wave run-up by about 70%. In contrast, Flow-3D™ was able to accurately simulate wave propagation, but under predicted wave generation by about 25% and over predicted the maximum wave run-up by about 40%. The question as to why both DualSPHysics and Flow-3D™ both over predict the maximum wave run-up during a SLGW simulation is still open. However, it is speculated that this due to a lack of either energy dissipation through air entrainment or eigenfrequency consideration’s.Applied Science, Faculty ofCivil Engineering, Department ofGraduat

State-of-the-Science on Postacute Rehabilitation: Setting a Research Agenda and Developing an Evidence Base for Practice and Public Policy. An Introduction

The Rehabilitation Research and Training Center on Measuring Rehabilitation Outcomes and Effectiveness, along with academic, professional, provider, accreditor, and other organizations, sponsored a 2-day State-of-the-Science of Postacute Rehabilitation Symposium in February 2007. The aim of this symposium was to serve as a catalyst for expanded research on postacute care (PAC) rehabilitation so that health policy is founded on a solid evidence base. The goals were to (a) describe the state of our knowledge regarding utilization, organization, and outcomes of postacute rehabilitation settings, (b) identify methodologic and measurement challenges to conducting research, (c) foster the exchange of ideas among researchers, policy makers, industry representatives, funding agency staff, consumers, and advocacy groups, and (d) identify critical questions related to setting, delivery, payment, and effectiveness of rehabilitation services. Plenary presentation and state-of-the-science summaries were organized around 4 themes: (a) the need for improved measurement of key rehabilitation variables and methods to collect and analyze this information, (b) factors that influence access to postacute rehabilitation care, (c) similarities and differences in quality and quantity of services across PAC settings, and (d) effectiveness of postacute rehabilitation services. The full set of symposium articles, including recommendations for future research, appear in Archives of Physical Medicine and Rehabilitation