The covalent and tertiary structure of bovine liver rhodanese

Bovine liver rhodanese is a single polypeptide of 293 amino acids in which the halves of the molecule assume analogous tertiary structures in the absence of substantial sequence homology. The sulphur atom transferred during catalysis is bound in persulphide linkage to Cys-247. Substrate binding seems to involve Arg-186 and Lys-249.     

Evolution of phosphofructokinase--gene duplication and creation of new effector sites

Phosphofructokinases (PFK; EC 2.7.1.11) are tetrameric enzymes that have a key role in the regulation of glycolysis; as such, they are subject to allosteric activation and inhibition by various metabolites. Eukaryotic PFKs are about twice the size of prokaryotic enzymes and are regulated by a wider repertoire of effectors: for example, the subunit molecular weights of rabbit muscle (RM) PFK and Bacillus stearothermophilus (Bs) PFK are 82,000 and 36,000, respectively. Both enzymes are activated by ADP (or AMP), but RM-PFK is also activated by fructose bisphosphates (FBP) and inhibited by ATP and citrate. This, together with other evidence, has led to speculation that mammalian PFKs have evolved by duplication of a prokaryotic gene, although previous peptide analysis failed to reveal internal homology in RM-PFK. Here we demonstrate clear homology among the N- and C-halves of RM-PFK and Bs-PFK, thus establishing an evolutionary relationship by series gene duplication and divergence. Furthermore, detailed knowledge of the Bs-PFK structure provides the basis for inferences concerning the structural organization of RM-PFK and the evolution of new effector sites in the enzyme tetramer.     

Membrane-anchored aspartyl protease with Alzheimer's disease beta-secretase activity

Mutations in the gene encoding the amyloid protein precursor (APP) cause autosomal dominant Alzheimer's disease. Cleavage of APP by unidentified proteases, referred to as beta- and gamma-secretases, generates the amyloid beta-peptide, the main component of the amyloid plaques found in Alzheimer's disease patients. The disease-causing mutations flank the protease cleavage sites in APP and facilitate its cleavage. Here we identify a new membrane-bound aspartyl protease (Asp2) with beta-secretase activity. The Asp2 gene is expressed widely in brain and other tissues. Decreasing the expression of Asp2 in cells reduces amyloid beta-peptide production and blocks the accumulation of the carboxy-terminal APP fragment that is created by beta-secretase cleavage. Solubilized Asp2 protein cleaves a synthetic APP peptide substrate at the beta-secretase site, and the rate of cleavage is increased tenfold by a mutation associated with early-onset Alzheimer's disease in Sweden. Thus, Asp2 is a new protein target for drugs that are designed to block the production of amyloid beta-peptide peptide and the consequent formation of amyloid plaque in Alzheimer's disease.