Replication of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) requires proteolytic processing of the replicase polyprotein by two viral cysteine proteases, a chymotrypsin-like protease (3CLpro) and a papain-like protease (PLpro). These proteases are important targets for development of antiviral drugs that would inhibit viral replication and reduce mortality associated with outbreaks of SARS-CoV. In this work, we describe the 1.85-Å crystal structure of the catalytic core of SARS-CoV PLpro and show that the overall architecture adopts a fold closely resembling that of known deubiquitinating enzymes. Key features, however, distinguish PLpro from characterized deubiquitinating enzymes, including an intact zinc-binding motif, an unobstructed catalytically competent active site, and the presence of an intriguing, ubiquitinlike N-terminal domain. To gain insight into the active-site recognition of the C-terminal tail of ubiquitin and the related LXGG motif, we propose a model of PLpro in complex with ubiquitinaldehyde that reveals well defined sites within the catalytic cleft that help to account for strict substrate-recognition motifs.membrane-associated protease ͉ ubiquitin-like domain

A new class of coordinatively unsaturated, monomeric organoscandium compounds, Cp*2Sc-R (Cp* = (rj5-C5Me5); R = halide, hydride, alkyl, alkenyl, alkynyl, aryl), has been prepared. Cp*2Sc-Cl is obtained from reaction of ScCl3(THF)3 (THF = tetrahydrofuran) with LiCp*, and Cp*2Sc-R (R = CH3, C6H5, C6H4CH3, CH2C6H5) from treatment of Cp*2Sc-Cl with the appropriate organoalkali reagent. These organoscandium compounds react with dihydrogen rapidly to yield R-H and Cp*2Sc-H. The tetrahydrofuran adducts Cp*2ScX(THF) (X = Cl, H, CH3) are obtained upon treatment of Cp*2Sc-X with tetrahydrofuran. Rapid exchange of dihydrogen with the hydride ligands of Cp*2Sc-H and Cp*2ScH(THF) occurs even at low temperatures. Other alkyl derivatives may be conveniently prepared by treatment of Cp*2Sc-H (or Cp*2ScH(THF)) with -olefins, e.g., Cp*2ScCH2CH3 from Cp*2Sc-H and ethylene. Aliene and Cp*2Sc-H afford Cp*2Sc(7j3-C3H5). Cp*2Sc-R (R = H, CH3, aryl) reacts with pyridine to yield Cp*2Sc(C, N-j;2-C5H4N), which crystallizes in the orthorhombic space group

Four isozymes of pyruvate kinase are differentially expressed in human tissue. Human pyruvate kinase isozyme M2 (hPKM2) is expressed in early fetal tissues and is progressively replaced by the other three isozymes, M1, R, and L, immediately after birth. In most cancer cells, hPKM2 is once again expressed to promote tumor cell proliferation. Because of its almost ubiquitous presence in cancer cells, hPKM2 has been designated as tumor specific PK-M2, and its presence in human plasma is currently being used as a molecular marker for the diagnosis of various cancers. The X-ray structure of human hPKM2 complexed with Mg(2+), K(+), the inhibitor oxalate, and the allosteric activator fructose 1,6-bisphosphate (FBP) has been determined to a resolution of 2.82 A. The active site of hPKM2 is in a partially closed conformation most likely resulting from a ligand-induced domain closure promoted by the binding of FBP. In all four subunits of the enzyme tetramer, a conserved water molecule is observed on the 2-si face of the prospective enolate and supports the hypothesis that a proton-relay system is acting as the proton donor of the reaction (1). Significant structural differences among the human M2, rabbit muscle M1, and the human R isozymes are observed, especially in the orientation of the FBP-activating loop, which is in a closed conformation when FBP is bound. The structural differences observed between the PK isozymes could potentially be exploited as unique structural templates for the design of allosteric drugs against the disease states associated with the various PK isozymes, especially cancer and nonspherocytic hemolytic anemia.

Phenylalanine hydroxylase converts phenylalanine to tyrosine, a rate-limiting step in phenylalanine catabolism and protein and neurotransmitter biosynthesis. It is tightly regulated by the substrates phenylalanine and tetrahydrobiopterin and by phosphorylation. We present the crystal structures of dephosphorylated and phosphorylated forms of a dimeric enzyme with catalytic and regulatory properties of the wild-type protein. The structures reveal a catalytic domain flexibly linked to a regulatory domain. The latter consists of an N-terminal autoregulatory sequence (containing Ser 16, which is the site of phosphorylation) that extends over the active site pocket, and an alpha-beta sandwich core that is, unexpectedly, structurally related to both pterin dehydratase and the regulatory domains of metabolic enzymes. Phosphorylation has no major structural effects in the absence of phenylalanine, suggesting that phenylalanine and phosphorylation act in concert to activate the enzyme through a combination of intrasteric and possibly allosteric mechanisms.