Establishing a Biochemical System for the Purification and ATPase activity of GST-Dbp5

The export of mRNA out of the nucleus is a crucial step for eukaryotic gene expression. The export of mRNA transcripts is aided by Mex67, which allows export through the nuclear pore complex doorways in the nuclear envelope. Once out of the nucleus, a protein known as Dbp5, bound to ATP, Gle1, and Nup42 aids in the directionality of mRNA export by helping remove Mex67 from the mRNA strand. Following interaction with RNA, Dbp5 then hydrolyzes ATP so that it unbinds the mRNA, allowing for enzyme recycling. Previous efforts worked towards the purification of Dbp5, but the attempts were unsuccessful due to low expression of recombinant protein in E.coli. In this project, I am focusing on enhancing the bacterial induction in order to establish robust purification of recombinant Dbp5. This will help in developing ATPase assays involving Dbp5, Nup42, and Gle1. These ATPase assays will aid in better understanding the effects of Nup42 and Gle1 on Dbp5’s ATPase activity and will allow for future study on Dbp5’s ATPase activity. In order to enhance the bacterial induction, E. coli cells were transformed with a GST-Dbp5 plasmid and were induced with varying amounts of IPTG to determine the best procedure for bacterial induction. Results from the bacterial induction have indicated that alternative methods for bacterial induction should be explored. Future experiments will look into further enhancing the bacterial induction of Dbp5 in order to establish a biochemical system analyzing the ATPase activity of GST-Dbp5

Multi-Isotope Geochemical Baseline Study of the Carbon Management Canada Research Institutes CCS Field Research Station (Alberta, Canada), Prior to CO2 Injection

Carbon capture and storage (CCS) is an industrial scale mitigation strategy for reducing anthropogenic CO2 from entering the atmosphere. However, for CCS to be routinely deployed, it is critical that the security of the stored CO2 can be verified and that unplanned migration from a storage site can be identified. A number of geochemical monitoring tools have been developed for this purpose, however, their effectiveness critically depends on robust geochemical baselines being established prior to CO2 injection. Here we present the first multi-well gas and groundwater characterisation of the geochemical baseline at the Carbon Management Canada Research Institutes Field Research Station. We find that all gases exhibit CO2 concentrations that are below 1%, implying that bulk gas monitoring may be an effective first step to identify CO2 migration. However, we also find that predominantly biogenic CH4 (∼90%–99%) is pervasive in both groundwater and gases within the shallow succession, which contain numerous coal seams. Hence, it is probable that any upwardly migrating CO2 could be absorbed onto the coal seams, displacing CH4. Importantly, 4He concentrations in all gas samples lie on a mixing line between the atmosphere and the elevated 4He concentration present in a hydrocarbon well sampled from a reservoir located below the Field Research Station (FRS) implying a diffusive or advective crustal flux of 4He at the site. In contrast, the measured 4He concentrations in shallow groundwaters at the site are much lower and may be explained by gas loss from the system or in situ production generated by radioactive decay of U and Th within the host rocks. Additionally, the injected CO2 is low in He, Ne and Ar concentrations, yet enriched in 84Kr and 132Xe relative to 36Ar, highlighting that inherent noble gas isotopic fingerprints could be effective as a distinct geochemical tracer of injected CO2 at the FRS

Establishing the geochemical baseline of the CMC FieldResearch Station, Alberta, Canada, prior to CO₂ injection andresolving the source and fate of natural CO₂ in the Morecambe and Rhyl Fields, East Irish Sea Basin, UK

Carbon capture and storage (CCS) is an industrial scale, cost-effective mitigation strategy to reduce anthropogenic CO₂ from entering the atmosphere. Successful deployment of CCS is critically dependent on the safe and secure storage of CO₂ injected into the subsurface for storage. Geochemical monitoring and modelling are an important tool in tracking the migration and fate of injected CO₂ throughout the entire life cycle of a CCS project, from baseline to post-closure. In this thesis, a wide range of geochemical tools are utilised to understand the geochemical baseline of the CMC Field Research Station, Alberta, Canada prior to CO₂ injection, and to identify the source and potential trapping mechanism of CO₂ residing within the Morecombe and Rhyl Fields within the East Irish Sea Basin. The CMC Research Institutes Field Research Station (FRS) is a purpose built test site for developing and demonstrating the monitoring of subsurface fluids following subsurface CO₂ injection. This thesis presents the first multi-well gas and groundwater characterisation of the geochemical baseline of the site. The work highlights that gases sampled from a range of depths exhibit low CO₂ concentrations, and that biogenic methane occurs pervasively in the shallow (<550 m) succession. Furthermore, the presence of a minor thermogenic component that increases with depth is also established, correlating with elevated levels of radiogenic 4He. 4He generation and expulsion has been mathematically modelled, highlighting that concentrations in some samples are above the level that could be generated from in-situ radioactive decay of U and Th within the FRS stratigraphy. This research shows that a resolvable radiogenic contribution to fluids and gases at the site, indicating a fluid connection to a deeper hydrocarbon producing formation in the Western Canadian Sedimentary Basin. The inherent geochemical fingerprints within the injected CO₂ were also determined, and found to be depleted in He, Ne and Ar, yet elevated in ⁸⁴Kr and ¹³²Xe relative to ³⁶Ar. This implies that inherent noble gas fingerprints could be used as an effective geochemical tracers of the injected CO₂ at this site. The Rhyl Field, located within the East Irish Sea Basin, is a producing gas field, with a recorded CO₂ concentration of 37%. Prior to this work the source of the CO₂ was unconstrained and no knowledge existed on the effect that this elevated concentration may have had on the mineralogy of the reservoir units. Identification of mantle derived noble gas isotopes confirms the presence of magmatic volatiles within the Rhyl field, however the stable isotopic profile of the produced gas does not exhibit a magmatic signature. The ratio of -36.2‰ suggests a highly organic source. Integrating the history of the basin with previously unpublished well reports and data relating to a hydrothermal event within the basin, allows the CO₂ source to be better constrained. Paleogene dyke emplacement likely heated coaliferous Type II/III source rock underlying the basin, causing the fractional generation of CH₄, CO₂ and N2 -providing the Rhyl field with its distinct bulk gas signature. Additionally, detailed mineralogical studies into the Rhyl field show no evidence of late stage CO₂- rock interaction resulting in the precipitation of known carbonate minerals which have previously been linked to CO₂ sequestration (including siderite, dolomite, calcite, dawsonite etc). Importantly, the Rhyl field was found to contain up to 0.22% tunisite (NaCa₂Al₄(CO₃)₄(OH)₈Cl) cement, a rare carbonate bearing mineral. Through studying formation water data, obtained from the exploration of hydrocarbons, the formation mechanism of tunisite has been constrained. This study highlights that this fluid chemistry, combined with extensive K-feldspar dissolution and CO₂ ingress in the early Paleogene, is the most likely means of tunisite precipitation in the Rhyl field. This work shows, for the first time, that tunisite can be a resolvable sink of CO₂ within the subsurface