Thesis (M.S.)--Wisconsin. / Includes bibliographical references (leaves 90-97).

Thesis (Ph.D.). / Written for the Dept. of Biochemistry. Title from title page of PDF (viewed 2009/06/11). Includes bibliographical references.

No description available.

Synechococcus, a genus of photosynthetic cyanobacteria, is the second most abundant oxygenic microorganism in the marine environment that contributes significantly to the ocean’s primary productivity (Humily et al. 2013; Shukla et al. 2012). They are capable of utilizing available light of different wavelengths in the visible spectrum to perform photosynthesis and fix carbon dioxide and thus inhabit a wide range of light niches in the ocean along horizontal (coast vs offshore) and vertical gradients (depth) (Humily et al. 2013). A gene encoding a putative lyase isomerase, mpeQ, is present in phycoerythrin-II encoding operon that is expressed constitutively and a gene encoding putative lyase, mpeW, is present in CA-4 genomic island whose expression is regulated by ambient light color were identified and characterized in Synechococcus sp. A15- 62, a strain having a blue light specialist phenotype in its basal state. The amino acid sequence of the proteins encoded by mpeW and mpeQ are similar to other characterized lyases and these genes are conserved in cyanobacteria strains containing the CA4-B genomic island, which controls CA4 (Humily et al. 2013). The MpeW and MpeQ proteins were produced in E. coli and co-expressed with recombinant HT-MpeA and phycoerythrobilin (PEB) synthesis machinery. Site directed mutants of the HT-MpeA protein (Cys75Ala, Cys83Ala, Cys140Ala) were used to investigate the site for bilin attachment. The recombinant protein co-expression experiments of MpeQ and MpeW demonstrated that MpeQ attaches phycoerythrobilin (PEB) to cysteine-83 site on a-phycoerythrin II and isomerizes it to phycourobilin (PUB) and MpeW attaches phycoerythrobilin (PEB) to the same site.

The biosynthesis of DMSP by phytoplankton and algae has wide ranging impact on marine organisms. Release of DMSP and uptake by marine bacteria leads to the eventual catabolism of this osmolyte. Enzymatic breakdown of DMSP leads to acrylate and volatile DMS production, which is fluxed into the atmosphere. When DMS enters the atmosphere it undergoes oxidation, acting as nucleation sites for water. The nucleation of water, and the subsequent cloud formation increases the albedo and reflects solar radiation. Global climate has therefore been hypothesized to be dependent upon DMSP breakdown to DMS. The enzymatic production of acrylate is also of interest for industrial applications. Only six enzymes are known to act as a DMSP-lyase, causing the production of DMS. These enzymes are still being discovered, and until recently there was very limited analysis of the biochemical requirements for catalysis. The work presented here investigates these requirements and the structural properties that permit the elimination reaction yielding DMS.

Thesis advisor: Thomas C. Chiles / B-lymphocytes respond to environmental cues for their survival, growth, and differentiation through receptor-mediated signaling pathways. Naïve Blymphocytes must acquire and metabolize external glucose in order to support the bioenergetics associated with maintaining cell volume, ion gradients, and basal macromolecular synthesis. The up-regulation of glycolytic enzyme expression and activity via engaged B-cell receptor mediated-events was glucose-dependent. This suggests an essential role for glucose energy metabolism in the promotion of B cell growth, survival, and proliferation in response to extracellular stimuli. In addition, the activity of ATP-citrate lyase (ACL) was determined to be crucial for ex vivo splenic B cell differentiation to antibody-producing cells wherein B cells undergo endomembrane synthesis and expansion. This investigation employed knockout murine models as well as chemical inhibitors to determine the signaling components and enzymes responsible for glucose utilization and incorporation into membrane lipids. These results point to a critical role for phosphatidylinositol 3- kinase (PI3K) in orchestrating cellular glucose energy metabolism and glucosedependent de novo lipogenesis for B lymphocyte responses. / Thesis (PhD) — Boston College, 2012. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Cupins are a large superfamily of enzymatic and non-enzymatic members that contain a conserved β-barrel domain, or double-stranded β-helix (DSBH) fold. The cupin superfamily is one of the most functionally diverse groups of proteins known to exist. The vast majority of cupins contain a mononuclear metal binding site at the core of the DSBH fold capable of binding different metal ions. One of the largest cupin subfamilies is known as the Fe(II)/α-ketoglutarate (αKG)-dependent dioxygenases. Prolyl 4-hydroxylases (P4Hs) belong to the group of Fe(II)/αKG-dependent dioxygenases and catalyze the formation of 4R-hydroxyproline (Hyp) from various proline-containing substrates. The formation of Hyp is an important post-translational modification to many different proteins involved in essential biochemical pathways. Abnormalities in these pathways can lead to diseases such as cancer, fibrosis, respiratory issues, scurvy, and stroke. An Fe(II)/αKG-dependent prolyl hydroxylase from Bacillus anthracis (BaP4H) was investigated to understand its substrate recognition ability and catalytic properties. Novel crystal structures were solved that revealed conformational changes upon substrate binding and key interactions of various ligands in the active site for different catalytic steps. Although the majority of cupin family enzymes catalyze a reaction using iron as an essential cofactor, other metal cofactors can allow the diverse biological transformations carried out by this group of enzymes. A class of enzymes known as dimethylsulfoniopropionate (DMSP) lyases uses different metal ions to catalyze the formation of acrylate and dimethylsulfide (DMS) from DMSP. DMSP is one of the most prevalent and significant molecules to the life and biogeochemistry of the oceans. The products DMS and acrylate are environmentally significant and industrially valuable. DMSP is predominantly catabolized by marine bacteria and can serve different functions. One of the most abundant bacteria in the ocean, Pelagibacter, was determined to contain a DMSP lyase DddK. The DddK catalyzed DMSP lyase activity in the presence of different metal ions has shown that it catalytically prefers Ni(II) compared to other transition metal ions examined. Spectroscopic, site-directed mutagenesis, and crystallographic studies illustrate central residues responsible for metal ion binding and possible roles in transition state stabilization. A greater mechanistic understanding of DMSP lyases will lead to more impactful information about global environmental climate regulation.

Universiẗat, Diss., 1997--Kaiserslautern.

Universiẗat, Diss., 2002--Potsdam.

Universiẗat, Diss., 2003--Münster (Westfalen).