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Journal Issue - Volume 14 Issue 12 (December 2005)

Abstract In the era of structural genomics, it is necessary to generate accurate structural alignments in order to build good templates for homology modeling. Although a great number of structural alignment algorithms have been developed, most of them ignore intermolecular interactions during the alignment procedure. Therefore, structures in different oligomeric states are barely distinguishable, and it is very challenging to find...

Abstract To obtain a gene construct for making single substitutions per channel and to determine the quaternary structure of the mechanosensitive channel MscL from Escherichia coli, covalent oligomers (monomer to hexamer) were engineered by gene fusion; up to six copies of the mscL gene were fused in tandem. All the multimeric tandem constructs yielded functional channels with wild‐type conductance and dwell times. Importantly, only the...

Abstract A sample of 35 independent molecular dynamics (MD) simulations of calmodulin (CaM) equilibrium dynamics was prepared from different but equally plausible initial conditions (20 simulations of the wild‐type protein and 15 simulations of the D129N mutant). CaM's radius of gyration and backbone mean‐square fluctuations were analyzed for the effect of the D129N mutation, and simulations were compared with experiments....

Abstract Many proteins that accumulate in the form of insoluble aggregates when they are overproduced in Escherichia coli can be rendered soluble by fusing them to E. coli maltose binding protein (MBP), and this will often enable them to fold in to their biologically active conformations. Yet, although it is an excellent solubility enhancer, MBP is not a particularly good affinity tag for protein purification. To compensate for this...

Abstract Structural genomics (SG) initiatives are expanding the universe of protein fold space by rapidly determining structures of proteins that were intentionally selected on the basis of low sequence similarity to proteins of known structure. Often these proteins have no associated biochemical or cellular functions. The SG success has resulted in an accelerated deposition of novel structures. In some cases the structural...

Abstract The structure of an AKAP docked to the dimerization/docking (D/D) domain of the type II (RIIα) isoform of protein kinase A (PKA) has been well characterized, but there currently is no detailed structural information of an AKAP docked to the type I (RIα) isoform. Dual‐specific AKAP2 (D‐AKAP2) binds in the nanomolar range to both isoforms and provided us with an opportunity to characterize the isoform‐selective nature of AKAP...

Abstract We describe in molecular detail how disruption of an intermonomer salt bridge (Arg337–Asp352) leads to partial destabilization of the p53 tetramerization domain and a dramatically increased propensity to form amyloid fibrils. At pH 4.0 and 37°C, a p53 tetramerization domain mutant (p53tet‐R337H), associated with adrenocortical carcinoma in children, readily formed amyloid fibrils, while the wild‐type (p53tet‐wt) did not. We...

Abstract The tryptophan repressor binding protein WrbA binds to the tryptophan repressor protein TrpR. Although the biological role of WrbA remains unclear, it has been proposed to function in enhancing the stability of TrpR–DNA complexes. Sequence database analysis has identified WrbA as a founding member of a flavodoxin‐like family of proteins. Here we present crystal structures of WrbA from Deinococcus radiodurans and Pseudomonas...

Abstract The phosphorylcholine esterase from Streptococcus pneumoniae, Pce, catalyzes the hydrolysis of phosphorylcholine residues from teichoic and lipoteichoic acids attached to the bacterial envelope and comprises a globular N‐terminal catalytic module containing a zinc binuclear center and an elongated C‐terminal choline‐binding module. The dependence of Pce activity on the metal/enzyme stoichiometry shows that the two equivalents of ...

Abstract The crystal structures of the Vβ17+ β chains of two human T cell receptors (TCRs), originally derived from the synovial fluid (SF4) and tissue (C5–1) of a patient with rheumatoid arthritis (RA), have been determined in native (SF4) and mutant (C5–1F104→Y/C187→S) forms, respectively. These TCR β chains form homo‐dimers in solution and in crystals. Structural comparison reveals that the main‐chain conformations in the CDR regions of the...

Abstract The overproduction of eukaryotic membrane proteins is a major impediment in their structural and functional characterization. Here we have used the nisin‐inducible expression system of Lactococcus lactis for the overproduction of 11 mitochondrial transport proteins from yeast. They were expressed at high levels in a functional state in the cytoplasmic membrane. The results also show that the level of expression is influenced by the...

Abstract Hemoglobins (Hbs) reversibly bind gaseous diatomic ligands (e.g., O2) as the sixth heme axial ligand of the penta‐coordinate deoxygenated form. Selected members of the Hb superfamily, however, display a functionally relevant hexa‐coordinate heme Fe atom in their deoxygenated state. Endogenous heme hexa‐coordination is generally provided in these Hbs by the E7 residue (often His), which thus modulates accessibility to the heme distal...

  • Evolution of an acylase active on cephalosporin C

  • Loredano Pollegioni, Simona Lorenzi, Elena Rosini, Giorgia Letizia Marcone, Gianluca Molla, Roberto Verga, Walter Cabri, Mirella S. Pilone
  • Published in Wiley Interscience on Jan 01, 2009
  • DOI: 10.1110/ps.051671705 (p 3064-3076)

Abstract Semisynthetic cephalosporins are synthesized from 7‐amino cephalosporanic acid, which is produced by chemical deacylation or by a two‐step enzymatic process of the natural antibiotic cephalosporin C. The known acylases take glutaryl‐7‐amino cephalosporanic acid as a primary substrate, and their specificity and activity are too low for cephalosporin C. Starting from a known glutaryl‐7‐amino cephalosporanic acid acylase as...

Abstract Bacterial response regulators are key regulatory proteins that function as the final elements of so‐called two‐component signaling systems. The activities of response regulators in vivo are modulated by phosphorylation that results from interactions between the response regulator and its cognate histidine protein kinase. The level of response regulator phosphorylation, which is regulated by intra‐or extracellular signals...

Abstract Ketol‐acid reductoisomerase (KARI; EC 1.1.1.86) catalyzes two steps in the biosynthesis of branched‐chain amino acids. Amino acid sequence comparisons across species reveal that there are two types of this enzyme: a short form (Class I) found in fungi and most bacteria, and a long form (Class II) typical of plants. Crystal structures of each have been reported previously. However, some bacteria such as Escherichia coli...

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