Author: Emily Van Uden
Publisher:
ISBN:
Category :
Languages : en
Pages : 272

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Author: Miklos Guttman
Publisher:
ISBN:
Category :
Languages : en
Pages : 184

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The LDLR family of receptors mediates the uptake of lipoprotein particles, and is essential for cholesterol homeostasis. The LDL receptor-related protein 1 (LRP-1) mediates internalization of a large number of diverse ligands and is widely implicated in Alzheimer's disease. Clusters of complement-type ligand binding repeats (CRs) in the LDL receptor family are thought to mediate the interactions between these receptors and their various ligands. Apolipoprotein E, a key ligand for cholesterol homeostasis, has been shown to interact with LDLR, LRP and VLDLR, through these clusters. LDLR and VLDLR each contain a single ligand-binding repeat cluster, whereas LRP contains three large clusters of ligand binding repeats, each with ligand binding functions. In order to study smaller units of these ligand binding clusters we have engineered a new approach to express and refold complement repeat (CR) domains in E. coli. This successfully produced high yields of refolded protein with the benefit of inexpensive isotope labeling for NMR studies. We have expressed a subdomain of sLRP3 (CR16-18) that has previously been shown to recapitulate ligand binding to the isolated receptor binding portion of ApoE (residues 130-149). Binding experiments with the ApoE recognition region of LDLR (LA3-5) and CR16-18 showed that each CR could interact with ApoE(130-149) and that a conserved W25/D30 pair within each repeat appears critical for high affinity. The triple repeat LA3-5 showed the expected interaction with the lipid complexed ApoE(1-191)*DMPC, but surprisingly CR16-18 did not interact with this form of ApoE. To understand these differences in ApoE binding affinity, we introduced mutations of conserved residues from LA5 into CR18, and produced a CR16-18 variant capable of binding ApoE(1-191)*DMPC. This change cannot fully be accounted for by the interaction with ApoE's proposed receptor binding region, therefore we speculate that LA5 is recognizing a distinct epitope on ApoE that may only exist in the lipid bound form. The combination of avidity effects with this distinct recognition process likely governs the ApoE-LDL receptor interaction. Since even the strongest interaction between ApoE(130-149) and a single repeat (CR17) was relatively weak, we constructed a CR17-ApoE(130-149) fusion protein to stabilize the interface for structural studies. The structure revealed a motif seen previously in all ligand CR interactions, in which lysine residues of the ligand interact with the calcium binding site of the CR. Like many ligands of CRs ApoE(130-149) binds as a helix, but with an unexpected turn at H140. These studies also revealed that little structural rearrangement occurs within CR17 upon binding. In addition, dynamics measurements of the free and bound CR17 reveal that certain regions become more ordered, while others become less ordered upon binding. The cytoplasmic tail of LRP, containing two NPXY motifs, has been implicated in the onset of Alzheimer's disease. To examine the intracellular interactions of LRP, as well as to separate which proteins bind to each NPXY motif and their phosphorylation dependence, each NPXY motif microdomain was prepared in both phosphorylated and non-phosphorylated forms and used to probe rodent brain extracts for binding proteins. Proteins that bound specifically to the microdomains were identified by LC-MS/MS, and confirmed by western blot. Recombinant proteins were then tested for binding to each NPXY motif. The NPXY450-- (membrane distal) was found to interact with a large number of proteins, many of which only bound the tyrosine-phosphorylated form. This microdomain also bound a significant number of other proteins in the unphosphorylated state. Many of the interactions were later confirmed to be direct with recombinant proteins. The NPXY44--3 (membrane proximal) bound many fewer proteins and only to the phosphorylated form.

Author: Robert Charles Kowal
Publisher:
ISBN:
Category :
Languages : en
Pages : 390

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Author: A.D. Roses
Publisher: Springer Science & Business Media
ISBN: 3642801099
Category : Medical
Languages : en
Pages : 208

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There is now considerable genetic evidence that the type 4 allele of the apolipoprotein E gene is a major susceptibility factor associated with late-onset Alzheimer's disease, the common form of the disease defined as starting after sixty years of age. The role of apolipoprotein E in normal brain metabolism and in the pathogenesis of Alzheimer's disease are new and exciting avenues of research. This book, written by the most outstanding scientists in this new filed, is the first presentation of results concerning the implications of apolipoprotein E on the genetics, cell biology, neuropathology, biochemistry, and therapeutic management of Alzheimer's disease.

Author: David Eun-Kwang Kang
Publisher:
ISBN:
Category :
Languages : en
Pages : 390

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Author: Johnny Eugene Croy
Publisher:
ISBN:
Category :
Languages : en
Pages : 390

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Author: Anothai Pocathikorn
Publisher:
ISBN:
Category : Lipoproteins
Languages : en
Pages : 302

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[Truncated abstract] The low density lipoprotein receptor-related protein (LRP), a member of the low-density lipoprotein (LDL) receptor gene family is involved in numerous biological processes including lipoprotein metabolism. This thesis concerns investigations into some aspects of LRP metabolism/regulation and possible roles in coronary artery disease (CAD). Specific aims were: to investigate the association between polymorphisms in the LRP gene and in its associated protein, the lipoprotein receptor-associated protein (RAP), with the risk of CAD; to extensively examine the influence of the LRP exon 22 C200T polymorphism on lipid metabolism; to develop and characterise assays for the mRNA expression of LRP and 2 other genes relevant to lipid metabolism, the LDL receptor (LDLR), and HMG CoA reductase (HMGCR); and finally, to apply the latter techniques to studies on the influence of genetic variation in LRP, and dietary and drug interventions, on LRP, LDLR and HMGCR mRNA expression in nucleated blood cells from healthy human subjects. Six hundred CAD subjects and 700 similarly aged controls were genotyped for 8 LRP gene polymorphisms as well as for the RAP V311M polymorphism. ... In the final phase of my studies, I examined the influence of 4 weeks therapy with a cholesterol lowering drug, an HMGCR inhibitor, atorvastatin (20mg daily), on the mRNA expression of LDLR, LRP and HMGCR in human nucleated blood cells. Twelve normal Caucasian male subjects aged 49 ± 5 (SD) years were studied. Plasma total cholesterol and LDL-C decreased by averages of 29 % and 41 % after the 4 week period. This was accompanied by an elevation in LDLR mRNA expression by approximately 30 35 %. In contrast, there was no significant effect on LRP and HMGCR mRNA expression. In conclusion, the original findings in this thesis included: demonstration of a strong influence of the LRP exon 22 C200T polymorphism on coronary artery disease and LDLR expression, but without a clear effect on fasting or postprandial lipid levels; data on the biological variation in LDLR and LRP gene expression in nucleated blood cells from normal subjects; the influence of an oral fat load on the expression viii of these genes, finding that LDLR was significantly depressed; and finally, the observation that statin therapy upregulated LDLR in nucleated blood cells.

Author: Neale Ridgway
Publisher: Elsevier
ISBN: 0444634495
Category : Science
Languages : en
Pages : 625

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Book Description
Biochemistry of Lipids: Lipoproteins and Membranes, Volume Six, contains concise chapters that cover a wide spectrum of topics in the field of lipid biochemistry and cell biology. It provides an important bridge between broad-based biochemistry textbooks and more technical research publications, offering cohesive, foundational information. It is a valuable tool for advanced graduate students and researchers who are interested in exploring lipid biology in more detail, and includes overviews of lipid biology in both prokaryotes and eukaryotes, while also providing fundamental background on the subsequent descriptions of fatty acid synthesis, desaturation and elongation, and the pathways that lead the synthesis of complex phospholipids, sphingolipids, and their structural variants. Also covered are sections on how bioactive lipids are involved in cell signaling with an emphasis on disease implications and pathological consequences. Serves as a general reference book for scientists studying lipids, lipoproteins and membranes and as an advanced and up-to-date textbook for teachers and students who are familiar with the basic concepts of lipid biochemistry References from current literature will be included in each chapter to facilitate more in-depth study Key concepts are supported by figures and models to improve reader understanding Chapters provide historical perspective and current analysis of each topic

Author: Larissa C. Wilsie
Publisher:
ISBN:
Category : Lipids
Languages : en
Pages : 136

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Author: James Whisstock
Publisher: Academic Press
ISBN: 0123864712
Category : Medical
Languages : en
Pages : 461

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Serpins are a group of proteins with similar structures that were first identified as a set of proteins able to inhibit proteases. The acronym serpin was originally coined because many serpins inhibit chymotrypsin-like serine proteases. This volume of Methods in Ezymology is split into 2 parts and comprehensively covers the subject.