Browsing by Subject "Specificity"
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Item Cognitive status and ra-marked referents of nominal expressions in Persian discourse(2014-05) Sadrai, MahmoudThis dissertation examines the referential status of the morpheme ra in Persian discourse. A nominal expression in Persian can occur with or without ra as illustrated below: a. nævar ra zæbt kærd tape ra record did `s/he recorded the tape' But not * `s/he recorded a tape' b. nævar zæbt kærd tape record did `s/he recorded a tape' But not * `s/he recorded the tape' a. nævar zæbt shod tape record became two possible interpretations: `the tape was recorded', or `a tape was recorded' But not b. * nævar ra zæbt shod. However, the exact nature of the conditions under which ra occurs is a source of disagreement in the literature on this morpheme. Various, and sometimes contradictory, assertions about the functions of this Persian morpheme have been made in the literature. While most researchers agree that 'ra' primarily marks direct objects (albeit there is evidence that it can also potentially mark other non-subjects), they differ in whether these nominal expressions are marked for definiteness (Phillott 1919, Lambton 1953, Ghomeshi 1997), or specificity (Browne 1970, Karimi 1989, Windfuhr 1990). In defining the referential status of ra-marked nominal expressions, I use a framework, the Givenness Hierarchy (Gundel, Hedberg, and Zacharski (1993)), that makes it possible to provide a more precise definition of `definiteness' and `specificity'. This framework allows for more precise and fine-grained primitive notions than the traditional concepts of specificity and definiteness. Distinctions in the Givenness Hierarchy are made in terms of cognitive statuses (the location of referents in the memory and attention of a hearer) at the point in the discourse just before a particular linguistic form is used. In this dissertation I examine naturally occurring data from forty (40) different sources, as well as constructed examples. The purpose for choosing a variety of sources of data for this study is to ensure that the results reflect language in use in a variety of genres and contexts. As such, I examine data that includes short stories, newspaper articles, children's stories, and a telephone conversation. The data examined contains a total of 17,611 words, yielding 408 instances of ra-marked DPs. Each expression containing `ra' is examined and coded for the highest cognitive status of the referent of the nominal expression. Using the cognitive statuses outlined in the Givenness Hierarchy, I examine the referent of each nominal expression containing `ra' to determine if it is type identifiable, referential, uniquely identifiable, familiar, activated, or in focus. I conclude that the referents of DPs marked with only ra are uniquely identifiable or higher. However, a more nuanced situation arises when ra co-occurs with another referential marker, namely -i (whose referential status remains unclear).Item Developing methods to understand and engineer protease cleavage specificity(2016-09) Lane, MichaelProteases are ubiquitous enzymes that comprise nearly 2% of all human genes. These robust enzymes are attractive potential therapeutics due to their catalytic turnover and capability for exquisite specificity. While most existing drugs require a stoichiometric ratio to function, therapeutic proteases could clear their targets much more efficiently. Unfortunately, existing technologies are inadequate for understanding and engineering therapeutic proteolytic specificities. My thesis work has focused on building the groundwork to enable these technologies to thrive. For the goal of engineering a new protease, it is currently necessary to identify prototype proteases for engineering efforts that have specificities similar to the desired target substrate. Current technologies are unable to characterize proteases adequately for this goal. Accordingly, I invested in developing a method for the accurate characterization of protease cleavage specificity. Our unique combination of mRNA display technology, Next-Generation Sequencing, and mass spectrometry enables the sampling of all possible permutations of octamer substrates and the identification of millions of cleavage sites. The throughput of our approach is orders of magnitude greater than the current state-of-the-art methods. The resulting high-resolution specificity maps can be applied to identify promising protease prototypes, predict human cross-reactivity, or lead to a better understanding of this critical component of natural physiology. In the work presented here, I applied my new specificity-screening method to assess the specificities of the proteases factor Xa, ADAM17, and streptopain. The resulting cleavage preference maps confirmed known specificities, and revealed new insight into the broad preferences of both narrow- and broad-specificity proteases. In particular, disfavored amino acids were illuminated better than ever before. The next focus of my work was to engineer multiple-subsite novel protease specificity. I chose streptopain as the prototype for my efforts to neutralize the superantigen exotoxin SpeA. I identified a target loop of SpeA wherein cleavage would result in inactive fragments. Further, I confirmed that streptopain can be successfully presented as an mRNA displayed fusion. In summary, my thesis work established crucial methodologies for applying mRNA display technology to enable the understanding and ultimately engineering the specificity of therapeutic proteases.