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NMR studies have long been used as a tool to derive structural and dynamic information. Such information has a wide range of applications, and notably is used in the study of structure-activity relationships. The aims of this work were to use NMR spectroscopy to derive structures of the molecules inhibiting the activation of the alternative pathway of the complement portion of the innate immune system (namely, the N-terminus of factor H (FH) and two small peptides, Compstatin 10 and Compstatin 20) and to consider the interdomain dynamics of proteins consisting of three modules theoretically (in silico) and experimentally (for the three N-terminal domains of FH). We focused on the three N-terminal complement control protein (CCP) domains of the important complement regulator, human factor H (i.e. FH1-3). Its three-dimensional solution structure was derived based on nuclear Overhauser effects and residual dipolar couplings (RDCs). Each of the three CCP modules in this structure was similar to the corresponding CCP in the previously derived C3b-bound structure of FH1-4, but the relative orientations of the domains were different. These orientations were additionally different from the interdomain orientations in other molecules that interact with C3b, such as DAF2-4 and CR1-15-17. The measured RDC datasets, collected under three different conditions in media containing magnetically aligned bicelles (disk-like particles formed from phospholipids), were used to estimate interdomain motions in FH1-3. A method in which the data was fitted to a structural ensemble was used to analyze such interdomain flexibility. More than 80% of the conformers of this predominantly extended three-domain molecule exhibit flexions of < 40°. Such segmental flexibility (together with the local dynamics of the hypervariable loop within domain 3) could facilitate recognition of C3b via initial anchoring, as well as eventual reorganization of modules into the conformation captured in the previously solved crystal structure of a C3b complex with FH1-4. The NMR study of the Compstatin analogues revealed unique structural features that had not before been observed in this group of peptides. These features included two b-turns per peptide, neither of which was located in the ‘canonical’ regions in which b-turns were observed in previous molecular dynamics and NMR studies. The structures of Compstatin 10 and Compstatin 20 derived here were consistent with the isothermal calorimetry (ITC) and surface plasmon resonance (SPR) data recorded previously. In the in silico study of interdomain motion of three-domain proteins carried out here, the domains were represented as vectors attached to one another in a linear fashion. They were allowed to undergo Brownian motion biased by the potentials between the sequential vectors. The resulting trajectories were analyzed using model-free and extended model-free formalism. The degree of coupling of the interdomain motion with overall motion was determined, along with a representation of the overall motion. The similarity between the trajectories of the vectors transformed to this overall motion frame and the results obtained from the model-free analysis was determined.

Adhesive bonding as an alternative method of joining materials together has many advantages over the more conventional joining methods such as fusion and spot welding, bolting and riveting. For example, adhesives can be used to bond dissimilar materials, adhesive joints have a high stiffness to weight ratio and the stress distribution within the joint is much improved. Stainless steels are commonly used in applications that would clearly benefit from adhesive bonding; architectural cladding, because of the large bond areas involved, and in the railway industry, due to improved acoustic insulation and greater fatigue resistance. The work presented in this thesis is concerned with adhesive bonding of stainless steels intended for structural applications. As a starting point to the investigation, a review of the literature was conducted, covering the intrinsic mechanisms of adhesion, the significance of the chemical and physical nature of the adherend surface, the types of structural adhesives, the methods of testing adhesive joints and surface characterisation techniques. The first experimental stage, involved a screening programme to evaluate a number of candidate adhesive systems and adherend surface pre-treatments. Standard single overlap shear and floating roller peel tests conducted in ambient conditions were employed in the discrimination and the degree of compatibility between adhesive and adherend, as measured by the proportion of cohesive failure on the post-fracture face, was also considered. In the second stage of the experimental work, lap shear tests were used to evaluate the affects of surface contamination on joint strength. In addition, lap shear and peel tests were considered to assess the significance of the adhesive bondline and primer thickness. In order to assess the environmental durability of adhesive joints, lap shear and peel tests were conducted after ageing in ambient and high humidity environments. To compliment the data, Boeing wedge crack extension tests were also carried out on adhesive bonded joints incorporating adherends with different surface conditions, to investigate the contribution to joint strength in ambient and adverse environments afforded by surface pre-treatment. The next stage of the experimental work was designed to evaluate the significance of the adherend type and its thickness on initial lap shear strength. Several different commercial grades and gauges of stainless steel were used in the tests, which were conducted at room temperature. The final stage of the experimental work was concentrated on the room temperature creep and dynamic fatigue performance of adhesive joints. Throughout the course of study a number of different surface analytical techniques were employed to physically and chemically characterise the surfaces of pm-bonded adherends and to identify the locus of failure on post-fracture faces. The single overlap shear and floating roller peel tests were able to differentiate between the candidate adhesives; epoxy systems, particularly the toughened variants, were considered the most suitable structural adhesives for bonding stainless steels in load bearing applications. However, these tests and subsequent tests using lap shear and peel, failed to discriminate conclusively between the different surface pre-treatments (except untreated or crudely prepared surfaces) and ageing environments. The Boeing wedge crack extension tests were found to be sensitive to the condition of the adherend surface and the environment in which the joint is located; roughening the surface of the adherend either chemically or physically was found to enhance joint durability in ambient, high humidity and sub-zero environments. The use of surface primers and coupling agents may protect the un-bonded surface and benefit joint durability, but excessively thick primer layers may reduce joint strength. The stiffness of the adherend material was found to significantly influence lap shear strength. Stiffer adherends, either thicker or inherently stronger, give higher joint strengths because they resist joint rotation and the peel stresses at the extremes of the overlap are minimised. Lap joints with low stiffness adherends will fail by peel-dominated, adherend-controlled failure and lap joints with high stiffness adherends will fail by shear-dominated, adhesive-controlled failure. Two elastic models were proposed for determining the elastic rotation and the line peel force as a function of the shear stress. The room temperature creep results showed an endurance limit of -40% mean static failure load (design load = 250 N.mni1 ). The dynamic fatigue results were favourable compared to those of spot welded and weldbonded joints and an endurance limit of 40% mean static failure load (design load = 250 N.mm-1 ) was observed. Finally, leaving the hard fillets of cured adhesive squeeze-out, intact at the extremes of the overlap, will reinforce the joint and minimise the rotation-induced peel stresses that will lead to premature failure when the adherend plastically deforms under static or dynamic loading.

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