Biomedical subjects

Daniel J Kenan

Publications and source records attributed to Daniel J Kenan.

8 recordsLinked to original sources

PubMed2006

Single-chain antibodies against DNA aptamers for use as adapter molecules on DNA tile arrays in nanoscale materials organization.

Complex DNA nanostructures have been developed as structural components for the construction of nanoscale objects. Recent advances have enabled self-assembly of organized DNA nanolattices and their use in patterning functional bio-macromolecules and other nanomaterials. Adapter molecules that bind specifically to both DNA lattices and nanomaterials would be useful components in a molecular construction kit for patterned nanodevices. Herein we describe the selection from phage display libraries of single-chain antibodies (scFv) for binding to a specific DNA aptamer and their development as adapter molecules for nanoscale construction. We demonstrate the decoration of various DNA tile structures with aptamers and show binding of the selected single-chain antibody as well as the self-assembly of mixed DNA-protein biomolecular lattices.

Antibodies↗

PubMed2006

Peptide-PEG amphiphiles as cytophobic coatings for mammalian and bacterial cells.

Amphiphilic macromolecules containing a polystyrene-adherent peptide domain and a cell-repellent poly(ethylene glycol) domain were designed, synthesized, and evaluated as a cytophobic surface coating. Such cytophobic, or cell-repellent, coatings are of interest for varied medical and biotechnological applications. The composition of the polystyrene binding peptide domain was identified using an M13 phage display library. ELISA and atomic force spectroscopy were used to evaluate the binding affinity of the amphiphile peptide domain to polystyrene. When coated onto polystyrene, the amphiphile reduced cell adhesion of two distinct mammalian cell lines and pathogenic Staphylococcus aureus strains.

Amino Acid Sequence↗

PubMed2006

Phage display screening for peptides that inhibit polyglutamine aggregation.

Proteins with expanded polyglutamine domains cause nine dominantly inherited, neurodegenerative diseases, including Huntington's disease. There are no therapies that inhibit disease onset or progression. To identify a novel therapeutic, we screened phage displayed peptide libraries for phage that bind preferentially to expanded polyglutamine repeats. We identified a peptide motif that inhibits polyglutamine aggregation in vitro and inhibits death in cellular and Drosophila models of the polyglutamine repeat diseases. In this chapter, we describe in detail how to screen a peptide phage display library and highlight results demonstrating the success of this approach. A similar experimental approach could be used for other diseases caused by conformational change in disease proteins, including prion, Alzheimer's, and Parkinson's diseases.

Amino Acid Sequence↗

PubMed2005

Angiostatin's molecular mechanism: aspects of specificity and regulation elucidated.

Tumor growth requires the development of new vessels that sprout from pre-existing normal vessels in a process known as "angiogenesis" [Folkman (1971) N Engl J Med 285:1182-1186]. These new vessels arise from local capillaries, arteries, and veins in response to the release of soluble growth factors from the tumor mass, enabling these tumors to grow beyond the diffusion-limited size of approximately 2 mm diameter. Angiostatin, a naturally occurring inhibitor of angiogenesis, was discovered based on its ability to block tumor growth in vivo by inhibiting the formation of new tumor blood vessels [O'Reilly et al. (1994a) Cold Spring Harb Symp Quant Biol 59:471-482]. Angiostatin is a proteolytically derived internal fragment of plasminogen and may contain various members of the five plasminogen "kringle" domains, depending on the exact sites of proteolysis. Different forms of angiostatin have measurably different activities, suggesting that much remains to be elucidated about angiostatin biology. A number of groups have sought to identify the native cell surface binding site(s) for angiostatin, resulting in at least five different binding sites proposed for angiostatin on the surface of endothelial cells (EC). This review will consider the data supporting all of the various reported angiostatin binding sites and will focus particular attention on the angiostatin binding protein identified by our group: F(1)F(O) ATP synthase. There have been several developments in the quest to elucidate the mechanism of action of angiostatin and the regulation of its receptor. The purpose of this review is to describe the highlights of research on the mechanism of action of angiostatin, its' interaction with ATP synthase on the EC surface, modulators of its activity, and issues that should be explored in future research related to angiostatin and other anti-angiogenic agents.

ATP Synthetase Complexes↗

PubMed2005

Ectopic localization of mitochondrial ATP synthase: a target for anti-angiogenesis intervention?

A receptor for angiostatin was identified on the surface of endothelial cells as F(1)-F(0) ATP synthase (Moser et al., 1999). Proc. Natl. Acad. Sci. U.S.A. 96, 2811-2816. This ectopic ATP synthase catalyzes ATP synthesis and is inhibited by angiostatin over a wide pH range. Endothelial cells grown at normal pH suffer no ill effects from this angiostatin-mediated inhibition of ATP synthase, whereas endothelial cells grown at low, tumor-like extracellular pH cannot maintain a normal intracellular pH and die. Angiostatin inhibits both ATP synthesis and ATP hydrolysis (Moser et al., 2001) and interferes with intracellular pH regulation (Wahl and Grant, 2002; Wahl et al., 2002). Although angiostatin administered intravenously is cleared from the circulation in a matter of minutes, angiostatin-mimetics that are more stable have potential for clinical application. An angiostatin-mimetic activity has recently been observed using a polyclonal antibody against the beta catalytic subunit of ATP synthase. In order to explore the mechanism of action of angiostatin and its mimetics, further work needs to be done to evaluate clinical applicability, specificity, and contraindications for this class of therapeutics.

Angiogenesis Inhibitors↗

PubMed2004

An Inhibitor of the F1 subunit of ATP synthase (IF1) modulates the activity of angiostatin on the endothelial cell surface.

Angiostatin binds to endothelial cell (EC) surface F(1)-F(0) ATP synthase, leading to inhibition of EC migration and proliferation during tumor angiogenesis. This has led to a search for angiostatin mimetics specific for this enzyme. A naturally occurring protein that binds to the F1 subunit of ATP synthase and blocks ATP hydrolysis in mitochondria is inhibitor of F1 (IF1). The present study explores the effect of IF1 on cell surface ATP synthase. IF1 protein bound to purified F(1) ATP synthase and inhibited F(1)-dependent ATP hydrolysis consistent with its reported activity in studies of mitochondria. Although exogenous IF1 did not inhibit ATP production on the surface of EC, it did conserve ATP on the cell surface, particularly at low extracellular pH. IF1 inhibited ATP hydrolysis but not ATP synthesis, in contrast to angiostatin, which inhibited both. In cell-based assays used to model angiogenesis in vitro, IF1 did not inhibit EC differentiation to form tubes and only slightly inhibited cell proliferation compared with angiostatin. From these data, we conclude that inhibition of ATP synthesis is necessary for an anti-angiogenic outcome in cell-based assays. We propose that IF1 is not an angiostatin mimetic, but it can serve a protective role for EC in the tumor microenvironment. This protection may be overridden in a concentration-dependent manner by angiostatin. In support of this hypothesis, we demonstrate that angiostatin blocks IF1 binding to ATP synthase and abolishes its ability to conserve ATP. These data suggest that there is a relationship between the binding sites of IF1 and angiostatin on ATP synthase and that IF1 could be employed to modulate angiogenesis.

Angiostatins↗

PubMed2004

La gets its wings.

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Autoantigens↗

PubMed2002

Direct interactions between corepressors and coactivators permit the integration of nuclear receptor-mediated repression and activation.

The unliganded thyroid hormone receptor beta (TRbeta) represses the basal transcriptional activity of target genes, in part through interactions with the nuclear receptor corepressor (N-CoR). In this study we have identified a rather unexpected interaction between N-CoR and the nuclear receptor coactivator ACTR. We have demonstrated in vitro and in intact cells that N-CoR directly associates with ACTR and that the interaction surfaces on N-CoR and ACTR are distinct from those required for TR binding. The significance of this finding was demonstrated by showing that N-CoR facilitates an interaction between unliganded-TRbeta and ACTR. One possible consequence of the formation of the trimeric complex of N-CoR/ACTR/unliganded-TR is that N-CoR may raise the local concentration of ACTR at target gene promoters. In support of this hypothesis it was demonstrated that the presence of N-CoR can enhance TRbeta-mediated transcriptional activation. It is proposed, therefore, that TRbeta- mediated activation and repression are integrally linked in a manner that is not predicted by the current models of nuclear receptor action.

Acetyltransferases↗