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The normal plasma protein serum amyloid P component (SAP) binds to fibrils in all types of amyloid deposits, and contributes to the pathogenesis of amyloidosis. In order to intervene in this process we have developed a drug, R-1-[6-[R-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl]pyrrolidine-2-carboxylic acid, that is a competitive inhibitor of SAP binding to amyloid fibrils. This palindromic compound also crosslinks and dimerizes SAP molecules, leading to their very rapid clearance by the liver, and thus produces a marked depletion of circulating human SAP. This mechanism of drug action potently removes SAP from human amyloid deposits in the tissues and may provide a new therapeutic approach to both systemic amyloidosis and diseases associated with local amyloid, including Alzheimer's disease and type 2 diabetes.

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The dynamic interplay between the extracellular matrix and embryonic stem cells (ESCs) constitutes one of the key steps in understanding stem cell differentiation in vitro. Here we report a biologically-active laminin-111 fragment generated by matrix metalloproteinase 2 (MMP2) processing, which is highly up-regulated during differentiation. We show that the β1-chain-derived fragment interacts via α3β1-integrins, thereby triggering the down-regulation of MMP2 in mouse and human ESCs. Additionally, the expression of MMP9 and E-cadherin is up-regulated in mouse ESCs-key players in the epithelial-to-mesenchymal transition. We also demonstrate that the fragment acts through the α3β1-integrin/extracellular matrix metalloproteinase inducer complex. This study reveals a previously unidentified role of laminin-111 in early stem cell differentiation that goes far beyond basement membrane assembly and a mechanism by which an MMP2-cleaved laminin fragment regulates the expression of E-cadherin, MMP2, and MMP9. L aminin-111, together with laminin-511, is among the first extracellular matrix (ECM) proteins expressed during early embryogenesis. Members of the laminin family are highly conserved between different species and are critical constituents of basement membranes in the early blastocyst as well as a variety of other tissues (1). The expression of the three laminin-111 chains-α1, β1, and γ1-is initiated as early as at the two-cell stage (2-4). Laminin-111-together with collagen IV, nidogen, perlecan, and other proteins-is assembled into the basement membrane (5), where it provides not only physical support, but also the capacity to modulate ECM function when modified by other proteins (6-8).The interactions between matrix metalloproteinases (MMPs) and ECM proteins have been extensively studied over the past decades (9, 10). The cellular expression of MMPs is precisely regulated such that ECM molecules are processed at different cell stages (11). It has been shown that laminin-111 can be processed by different MMPs and a variety of other enzymes and that such modifications are mainly related to cell migration because cleavage of laminin results in a loosened basement membrane (6,8,12,13). Recent insights into the existence of "cryptic" ECM interaction sites-sequences usually hidden within the tertiary structure of the protein or within the assembly of the ECM-have enabled a greater understanding of how cells interact with the ECM and of the astonishingly complex interaction between cells and ECM proteins (14). The ECM can no longer be thought of as just a passive scaffold and physical support for cells in vivo or as a more-or-less adequate cell culture substrate in vitro; the ECM and its modification is now known to act as one of the key constituents in cell regulation. Therefore, to successfully model development in vitro by using pluripotent stem cells, it is imperative to give careful consideration to cell-ECM interactions.In this work, we address the question of whether MMP2 (15-17) modifies laminin-111 and whether s...

The selective binding of serum amyloid P component (SAP) to proteins in the pathological amyloid cross-L L fold suggests a possible chaperone role. Here we show that human SAP enhances the refolding yield of denatured lactate dehydrogenase and protects against enzyme inactivation during agitation of dilute solutions. These effects are independent of calcium ions and are not inhibited by compounds that block the amyloid recognition site on the B face of SAP, implicating the A face and/or the edges of the SAP pentamer. We discuss the possibility that the chaperone property of SAP, or its failure, may contribute to the pathogenesis of amyloidosis.z 2000 Federation of European Biochemical Societies.

Background: Laminin self-assembly into a cell-associated network is essential for basement membrane formation.Results: The isolated tips of the laminin short arms form ternary complexes in solution.Conclusion: The nodes in the laminin network are formed by the N-terminal domains of one α, one β, and one γ chain.Significance: The reconstitution of laminin network nodes enables structure-function studies.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. This license does not permit commercial exploitation or the creation of derivative works without specific permission.The CBF3 complex is an essential core component of the budding yeast kinetochore and is required for the centromeric localization of all other kinetochore proteins. We determined the crystal structure of a large section of the protein Cep3 from CBF3, which is the only component with obvious DNA-binding motifs. The protein adopts a roughly bilobal shape, with an extended dimerization interface. The dimer has a large central channel that is sufficient to accommodate duplex B-form DNA. The zincfinger domains emerge at the edges of the channel, and could bind to the DNA in a pseudo-symmetrical manner at degenerate half-sites in the centromeric sequence. We propose a mechanism for the modulation of DNA affinity by an acidic activator domain, which could be applicable to a wider family of transcription factors.

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