Louise H. Foley

Regulation of Tiam1 Nucleotide Exchange Activity by Pleckstrin Domain Binding Ligands

Rho family GTPases play roles in cytoskeletal organization and cellular transformation. Tiam1 is a member of the Dbl family of guanine nucleotide exchange factors that activate Rho family GTPases. These exchange factors have in common a catalytic Dbl homology and adjacent pleckstrin homology domain. Previous structural studies suggest that the pleckstrin domain, a putative phosphoinositide-binding site, may serve a regulatory function. We identified ascorbyl stearate as a compound that binds to the pleckstrin domain of p120 Ras GTPase-activating protein. Furthermore, ascorbyl stearate appears to be a general pleckstrin domain ligand, perhaps by mimicking an endogenous amphiphilic ligand. Tiam1 nucleotide exchange activity was greatly stimulated by ascorbyl stearate. Certain phosphoinositides also stimulated Tiam1 activity but were less potent than ascorbyl stearate. Tiam1 contains an additional N-terminal pleckstrin domain, but only the C-terminal pleckstrin domain was required for activation. Our results suggest that the pleckstrin domains of Dbl-type proteins may not only be involved in subcellular localization but may also directly regulate the nucleotide exchange activity of an associated Dbl homology domain. In addition, this paper introduces ascorbyl stearate as a pleckstrin domain ligand that can modulate the activity of certain pleckstrin domain-containing proteins.

X-ray structure of a novel matrix metalloproteinase inhibitor complexed to stromelysin.

A new class of matrix metalloproteinase (MMP) inhibitors has been identified by screening a collection of compounds against stromelysin. The inhibitors, 2,4,6‐pyrimidine triones, have proven to be potent inhibitors of gelatinases A and B. An X‐ray crystal structure of one representative compound bound to the catalytic domain of stromelysin shows that the compounds bind at the active site and ligand the active‐site zinc. The pyrimidine triones mimic substrates in forming hydrogen bonds to key residues in the active site, and provide opportunities for placing appropriately chosen groups into the S1′ specificity pocket of MMPs. A number of compounds have been synthesized and assayed against stromelysin, and the variations in potency are explained in terms of the binding mode revealed in the X‐ray crystal structure.

Novel 5,5-disubstitutedpyrimidine-2,4,6-triones as selective MMP inhibitors

The 5,5-disubstitutedpyrimidine-2,4,6-triones represent a new class of MMP inhibitors showing selectivity for the gelatinases A and B, collagenase-3, and human neutrophil collagenase. The SAR presented here is in good agreement with an X-ray structure of compound 5 bound to the catalytic domain of stromelysin-1. While of the barbiturate structural class, compound 5 did not show any toxic or sedative effects.

Modified 3-alkyl-1,8-dibenzylxanthines as GTP-competitive inhibitors of phosphoenolpyruvate carboxykinase

The first non-substrate like inhibitors of human cytosolic phosphoenolpyruvate carboxykinase (PEPCK) competitive with GTP are reported. An effort to discover orally active compounds that improve glucose homeostasis in Type 2 diabetics by reversibly inhibiting PEPCK led to the discovery of 1-allyl-3-butyl-8-methylxanthine (5). We now report modifications at N-1 and C-8 that improved the in vitro activity of the initial xanthine HTS hit by 100-fold and a developing SAR for this class of inhibitor.

X-ray structures of two xanthine inhibitors bound to PEPCK and N-3 modifications of substituted 1,8-dibenzylxanthines.

The analysis of the X-ray structures of two xanthine inhibitors bound to PEPCK and a comparison to the X-ray structure of GTP bound to PEPCK are reported. The SAR at N-1, N-7 and developing SAR at C-8 are consistent with information gained from the X-ray structures of compounds 1 and 2 bound to PEPCK. Representative N-3 modifications of compound 2 that led to the discovery of 3-cyclopropylmethyl and its carboxy analogue as optimal N-3 groups are presented.

Ascorbic Acid Chemistry. A Novel LiAlH4 Reduction Product and its Rearrangement

Abstract Reaction of 1 with LiAlH4 affords the chiral monobenzyloxy diol, 5(S), 6-0-isopropylidene-4(R)-hydroxyl-2-benzyloxy-2-hexenol. Evidence is provided to support a mechanism involving Michael addition of hydride to C-3. Reduction products, 2 & 3, undergo facile allylic alcohol rearrangements.