Kim Jonelle Stutzman-Engwall
Pfizer
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Publication
Featured researches published by Kim Jonelle Stutzman-Engwall.
Nature Structural & Molecular Biology | 2007
David Cunningham; Dennis E. Danley; Kieran F. Geoghegan; Matthew C. Griffor; Julie Hawkins; Timothy A. Subashi; Alison H. Varghese; Mark Ammirati; Jeffrey S. Culp; Lise R. Hoth; Mahmoud N. Mansour; Katherine M McGrath; Andrew P. Seddon; Shirish Shenolikar; Kim Jonelle Stutzman-Engwall; Laurie C. Warren; Donghui Xia; Xiayang Qiu
Proprotein convertase subtilisin kexin type 9 (PCSK9) lowers the abundance of surface low-density lipoprotein (LDL) receptor through an undefined mechanism. The structure of human PCSK9 shows the subtilisin-like catalytic site blocked by the prodomain in a noncovalent complex and inaccessible to exogenous ligands, and that the C-terminal domain has a novel fold. Biosensor studies show that PCSK9 binds the extracellular domain of LDL receptor with Kd = 170 nM at the neutral pH of plasma, but with a Kd as low as 1 nM at the acidic pH of endosomes. The D374Y gain-of-function mutant, associated with hypercholesterolemia and early-onset cardiovascular disease, binds the receptor 25 times more tightly than wild-type PCSK9 at neutral pH and remains exclusively in a high-affinity complex at the acidic pH. PCSK9 may diminish LDL receptors by a mechanism that requires direct binding but not necessarily receptor proteolysis.
Nature Structural & Molecular Biology | 2007
Xiayang Qiu; Anil Mistry; Mark Ammirati; Boris A. Chrunyk; Ronald W. Clark; Yang Cong; Jeffrey S. Culp; Dennis E. Danley; Thomas B. Freeman; Kieran F. Geoghegan; Matthew C. Griffor; Steven J. Hawrylik; Cheryl Myers Hayward; Preston Hensley; Lise R. Hoth; George A. Karam; Maruja E. Lira; David B. Lloyd; Katherine M McGrath; Kim Jonelle Stutzman-Engwall; Ann Subashi; Timothy A. Subashi; John F. Thompson; Ing-Kae Wang; Honglei Zhao; Andrew P. Seddon
Cholesteryl ester transfer protein (CETP) shuttles various lipids between lipoproteins, resulting in the net transfer of cholesteryl esters from atheroprotective, high-density lipoproteins (HDL) to atherogenic, lower-density species. Inhibition of CETP raises HDL cholesterol and may potentially be used to treat cardiovascular disease. Here we describe the structure of CETP at 2.2-Å resolution, revealing a 60-Å-long tunnel filled with two hydrophobic cholesteryl esters and plugged by an amphiphilic phosphatidylcholine at each end. The two tunnel openings are large enough to allow lipid access, which is aided by a flexible helix and possibly also by a mobile flap. The curvature of the concave surface of CETP matches the radius of curvature of HDL particles, and potential conformational changes may occur to accommodate larger lipoprotein particles. Point mutations blocking the middle of the tunnel abolish lipid-transfer activities, suggesting that neutral lipids pass through this continuous tunnel.
Protein Expression and Purification | 2009
David John Wasilko; S. Edward Lee; Kim Jonelle Stutzman-Engwall; Beverly A. Reitz; Thomas L. Emmons; Karl J. Mathis; Michael J. Bienkowski; Alfredo G. Tomasselli; H. David Fischer
Compounds capable of stimulating soluble guanylate cyclase (sGC) activity might become important new tools to treat hypertension. While rational design of these drugs would be aided by elucidation of the sGC three-dimensional structure and molecular mechanism of activation, such efforts also require quantities of high quality enzyme that are challenging to produce. We implemented the titerless infected-cells preservation and scale-up (TIPS) methodology to express the heterodimeric sGC. In the TIPS method, small-scale insect cell cultures were first incubated with a recombinant baculovirus which replicated in the cells. The baculovirus-infected insect cells (BIIC) were harvested and frozen prior to cell lysis and the subsequent escape of the newly replicated virus into the culture supernatant. Thawed BIIC stocks were ultimately used for subsequent scale up. As little as 1 mL of BIIC was needed to infect a 100-L insect cell culture, in contrast to the usual 1L of high-titer, virus stock supernatants. The TIPS method eliminates the need and protracted time for titering virus supernatants, and provides stable, concentrated storage of recombinant baculovirus in the form of infected cells. The latter is particularly advantageous for virus stocks which are unstable, such as those for sGC, and provides a highly efficient alternative for baculovirus storage and expression. The TIPS process enabled efficient scale up to 100-L batches, each producing about 200mg of active sGC. Careful adjustment of expression culture conditions over the course of several 100-L runs provided uniform starting titers, specific activity, and composition of contaminating proteins that facilitated development of a process that reproducibly yielded highly active, purified sGC.
Organic and Biomolecular Chemistry | 2003
Sabine Gaisser; Laurenz Kellenberger; Andrew L. Kaja; Alison J. Weston; Rachel E. Lill; Gabriele Wirtz; Steven Gary Kendrew; Lindsey Low; Rose M. Sheridan; Barrie Wilkinson; Ian S. Galloway; Kim Jonelle Stutzman-Engwall; Hamish McArthur; James Staunton; Peter F. Leadlay
Ivermectin, a mixture of 22,23-dihydroavermectin B1a9 with minor amounts of 22,23-dihydroavermectin B1b 10, is one of the most successful veterinary antiparasitic drugs ever produced. In humans, ivermectin has been used for the treatment of African river blindness (onchocerciasis) resulting in an encouraging decrease in the prevalence of skin and eye diseases linked to this infection. The components of ivermectin are currently synthesized by chemical hydrogenation of a specific double bond at C22-C23 in the polyketide macrolides avermectins B1a 5 and B1b 6, broad-spectrum antiparasitic agents isolated from the soil bacterium Streptomyces avermitilis. We describe here the production of such compounds (22,23-dihydroavermectins B1a 9 and A1a 11) by direct fermentation of a recombinant strain of S. avermitilis containing an appropriately-engineered polyketide synthase (PKS). This suggests the feasibility of a direct biological route to this valuable drug.
Microbiology | 1994
Meirwyn Evans; Frank S. Kaczmarek; Kim Jonelle Stutzman-Engwall; Paul Dyson
The degradation of Streptomyces avermitilis DNA samples analysed by conventional pulsed-field gel electrophoresis was shown to be due to Tris-dependent, double-strand cleavage. Using alternative electrophoretic conditions, separation of intact DNA molecules was achieved, permitting the identification of two novel giant linear plasmids: the 100 kb pSA1 and 250 kb pSA2. Use of pSA2 DNA as a probe showed that pSA1 does not cross-hybridize, indicating that the plasmids are not closely related. The site-specificity of the DNA modifications, which render the DNA susceptible to Tris-dependent cleavage, was found to be essentially identical to that of similar modifications found in the DNA of S. lividans.
Metabolic Engineering | 2005
Kim Jonelle Stutzman-Engwall; Steve Conlon; Ronald Fedechko; Hamish McArthur; Katja Pekrun; Yan Chen; Stephane J. Jenne; Charlene La; Na Trinh; Seran Kim; Ying-Xin Zhang; Richard J. Fox; Claes Gustafsson; Anke Krebber
Biotechnology and Bioengineering | 2003
Kim Jonelle Stutzman-Engwall; Steve Conlon; Ronald Fedechko; Frank S. Kaczmarek; Hamish McArthur; Anke Krebber; Yan Chen; Jeremy Minshull; Sun Ai Raillard; Claes Gustafsson
Biotechnology and Bioengineering | 2005
Gargi Seth; Robin Philp; Claudio D. Denoya; Katherine M McGrath; Kim Jonelle Stutzman-Engwall; Miranda Yap; Wei Shou Hu
Archive | 1995
Claudio D. Denoya; Kim Jonelle Stutzman-Engwall
Archive | 2003
Kim Jonelle Stutzman-Engwall; Anke Krebber; Claes Gustafsson; Jeremy Minshull; Sun Ai Raillard; Seran Kim; Yan Chen