Role of divalent cations in the 3',5'-exonuclease reaction of DNA polymerase I.


:X-ray studies of the proofreading 3',5'-exonuclease site of the large (Klenow) fragment of DNA polymerase I have detected a binuclear metal complex consisting of a pentacoordinate metal (site A) which shares a ligand, Asp-355, with an octahedral metal (site B) [Freemont, P. S., Friedman, J. M., Beese, L. S., Sanderson, M. R., & Steitz, T. A. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 8924-8928; Beese, L. S., & Steitz, T. A. (1991) EMBO J. 10, 25-33]. Kinetic studies of the activation of the 3',5'-exonuclease reaction by Co2+, Mn2+, or Mg2+, at low concentrations of DNA, reveal sigmoidal activation curves for the three metal ions with Hill coefficients of 2.3-2.4 and K0.5 values of 16.6 microM, 4.2 microM, and 343 microM, respectively. The binding of Co2+ to the enzyme results in the appearance of an intense visible absorption spectrum of the metal ion with maxima at 633, 570, and 524 nm and extinction coefficients of 190, 194, and 150 M-1 cm-1, respectively, suggesting the formation of a pentacoordinate Co2+ complex. Optical titration with Co2+ yields a sigmoidal titration curve which is best fit by assuming the cooperative binding of three Co2+ ions with a K0.5 of 39.9 microM, comparable to the value of 16.6 microM obtained kinetically. Displacement of Co2+ by 1 equiv of Zn2+, which binds tightly to the A site of the 3',5'-exonuclease, shifts the optical spectrum to 524 nm and lowers the extinction coefficient to 30 -1 cm-1, indicative of octahedral coordination.2+ the formation of the binuclear complex.






Han H,Rifkind JM,Mildvan AS




Has Abstract


1991-11-19 00:00:00












  • Quantification of cysteinyl S-nitrosylation by fluorescence in unbiased proteomic studies.

    abstract::Cysteinyl S-nitrosylation has emerged as an important post-translational modification affecting protein function in health and disease. Great emphasis has been placed on global, unbiased quantification of S-nitrosylated proteins because of physiologic and oxidative stimuli. However, current strategies have been hamper...


    pub_type: 杂志文章


    authors: Wiktorowicz JE,Stafford S,Rea H,Urvil P,Soman K,Kurosky A,Perez-Polo JR,Savidge TC

    更新日期:2011-06-28 00:00:00

  • Phosphorylation of c-Abl by protein kinase Pak2 regulates differential binding of ABI2 and CRK.

    abstract::The tyrosine kinase c-Abl is implicated in a variety of cellular processes that are tightly regulated by c-Abl kinase activity and/or by interactions between c-Abl and other signaling molecules. The interaction of c-Abl with the Abl interactor protein Abi2 is shown to be negatively regulated by phosphorylation of seri...


    pub_type: 杂志文章


    authors: Jung JH,Pendergast AM,Zipfel PA,Traugh JA

    更新日期:2008-01-22 00:00:00

  • Structure and function of DNA photolyase.

    abstract::Cyclobutane pyrimidine dimers (Pyr < > Pyr) are the major DNA photoproducts induced by the UV component of solar radiation. Photoreactivating enzyme (DNA photolyase) repairs DNA by utilizing the energy of visible light to break the cyclobutane ring of the dimer. Photolyases are monomeric proteins of 50-60 kDa with sto...


    pub_type: 杂志文章,评审


    authors: Sancar A

    更新日期:1994-01-11 00:00:00

  • Stable substructures of eightfold beta alpha-barrel proteins: fragment complementation of phosphoribosylanthranilate isomerase.

    abstract::The (beta alpha)8 (or "TIM")-barrel protein phosphoribosylanthranilate isomerase from Saccharomyces cerevisiae was cleaved between the sixth and seventh beta alpha module to test the capacity of the resulting fragments to adopt native format autonomously. The fragments, which were expressed from separate coding sequen...


    pub_type: 杂志文章


    authors: Eder J,Kirschner K

    更新日期:1992-04-14 00:00:00

  • Crystal Structures of Apo and Liganded 4-Oxalocrotonate Decarboxylase Uncover a Structural Basis for the Metal-Assisted Decarboxylation of a Vinylogous β-Keto Acid.

    abstract::The enzymes in the catechol meta-fission pathway have been studied for more than 50 years in several species of bacteria capable of degrading a number of aromatic compounds. In a related pathway, naphthalene, a toxic polycyclic aromatic hydrocarbon, is fully degraded to intermediates of the tricarboxylic acid cycle by...


    pub_type: 杂志文章


    authors: Guimarães SL,Coitinho JB,Costa DM,Araújo SS,Whitman CP,Nagem RA

    更新日期:2016-05-10 00:00:00

  • NMR studies of a deoxyribodecanucleotide containing an extrahelical thymidine surrounded by an oligo(dA).oligo(dT) tract.

    abstract::One- and two-dimensional NMR experiments were carried out on a decamer, d-(CGCTTTTCGC).d(GCGAAAAGCG), and on the same sequence with the addition of an unpaired thymidine, d(CGCTTTTCGC).d(GCGAATAAGCG), which will be referred to as the T-bulge decamer. Evidence from one-dimensional NOE experiments on the exchangeable pr...


    pub_type: 杂志文章


    authors: Morden KM,Gunn BM,Maskos K

    更新日期:1990-09-18 00:00:00

  • Key role of phenylalanine 20 in cytochrome c3: structure, stability, and function studies.

    abstract::Aromatic residues in c-type cytochromes might have an important function in the folding and/or electron transferring properties of the molecule. In the tetraheme cytochrome c3 (Mr 13 000) from Desulfovibrio vulgaris Hildenborough, Phe20, is located between heme 1 and heme 3 with its aromatic ring close and almost para...


    pub_type: 杂志文章


    authors: Dolla A,Arnoux P,Protasevich I,Lobachov V,Brugna M,Giudici-Orticoni MT,Haser R,Czjzek M,Makarov A,Bruschi M

    更新日期:1999-01-05 00:00:00

  • Thermodynamics of ion-induced RNA folding in the hammerhead ribozyme: an isothermal titration calorimetric study.

    abstract::The hammerhead ribozyme undergoes a well-defined two-stage conformational folding process, induced by the binding of magnesium ions. In this study, we have used isothermal titration calorimetry to analyze the thermodynamics of magnesium binding and magnesium ion-induced folding of the ribozyme. Binding to the natural ...


    pub_type: 杂志文章


    authors: Hammann C,Cooper A,Lilley DM

    更新日期:2001-02-06 00:00:00

  • Streptococcus pneumoniae isoprenoid biosynthesis is downregulated by diphosphomevalonate: an antimicrobial target.

    abstract::The toll that Streptococcus pneumoniae exacts on the welfare of humanity is enormous. This organism claims the lives of approximately 3700 people daily, the majority of whom are children below the age of 5, and the situation could worsen due to the increasing incidence of pernicious, multiple-antibiotic-resistant stra...


    pub_type: 杂志文章


    authors: Andreassi JL 2nd,Dabovic K,Leyh TS

    更新日期:2004-12-28 00:00:00

  • Methionine sulfoxide cytochrome c.

    abstract::Cytochrome c has been chemically modified by methylene blue mediated photooxidation. It is established that the methionine residues of the protein have been specifically converted to methionine sulfoxide residues. No oxidation of any other amino acid residues or the cysteine thioether bridges of the molecule occurs du...


    pub_type: 杂志文章


    authors: Ivanetich KM,Bradshaw JJ,Kaminsky LS

    更新日期:1976-03-09 00:00:00

  • Cholesterol-mediated conformational changes in the steroidogenic acute regulatory protein are essential for steroidogenesis.

    abstract::Although the mechanism by which the steroidogenic acute regulatory protein (StAR) promotes steroidogenesis has been studied extensively, it remains incompletely characterized. Because structural analysis has revealed a hydrophobic sterol-binding pocket (SBP) within StAR, this study sought to examine the regulatory rol...


    pub_type: 杂志文章


    authors: Rajapaksha M,Kaur J,Bose M,Whittal RM,Bose HS

    更新日期:2013-10-15 00:00:00

  • Linkage and sequence analysis of mannose-rich glycoprotein core oligosaccharides by proton nuclear magnetic resonance spectroscopy.

    abstract::The anomeric proton (H-1) chemical shifts of D-mannopyranosides in aqueous solution are affected both by the aglycon and by substitution of the ring [Lee, Y. C., & Ballou, C. E. (1965) Biochemistry 4, 257]. We have examined the 1H NMR spectra for a variety of linear and branched mannooligosaccharides and have assigned...


    pub_type: 杂志文章


    authors: Cohen RE,Ballou CE

    更新日期:1980-09-02 00:00:00

  • The C-terminal aqueous-exposed domain of the 45 kDa subunit of the particulate methane monooxygenase in Methylococcus capsulatus (Bath) is a Cu(I) sponge.

    abstract::The crystal structure of the particulate methane monooxygenase (pMMO) from Methylococcus capsulatus (Bath) has been reported recently [Lieberman, R. L., and Rosenzweig, A. C. (2005) Crystal structure of a membrane-bound metalloenzyme that catalyses the biological oxidation of methane, Nature 434, 177-182]. Subsequent ...


    pub_type: 杂志文章


    authors: Yu SS,Ji CZ,Wu YP,Lee TL,Lai CH,Lin SC,Yang ZL,Wang VC,Chen KH,Chan SI

    更新日期:2007-12-04 00:00:00

  • Mechanism of 25-hydroxyvitamin D3 24-hydroxylation: incorporation of oxygen-18 into the 24 position of 25-hydroxyvitamin D3.

    abstract::The oxygen enzymatically inserted as a hydroxyl function by chick kidney homogenate into the 24 position of 25-hydroxyvitamin D3 to give 24,25-dihydroxyvitamin D3 is derived exclusively from 18O2. Therefore, like the 25-hydroxyvitamin D3-lalpha-hydroxylase system, the 25-hydroxyvitamin D3-24-hydroxylase system is also...


    pub_type: 杂志文章


    authors: Madhok TC,Schnoes HK,DeLuca HF

    更新日期:1977-05-17 00:00:00

  • Conformational drift and cryoinactivation of lactate dehydrogenase.

    abstract::Solutions of porcine lactate dehydrogenase of micromolar concentration kept at 4 degrees C for several days lose the greater part of their enzymic activity but recover it when returned to room temperature. The rate of spoiling decreases and the rate of recovery increases with the concentration of the solutions. The de...


    pub_type: 杂志文章


    authors: King L,Weber G

    更新日期:1986-06-17 00:00:00

  • Electric field induced transient pores in phospholipid bilayer vesicles.

    abstract::A study of the voltage induction of transient pores in phospholipid bilayer vesicles is reported. Unilamellar vesicles (dipalmitoylphosphatidylcholine), with a size distribution of 100 +/- 30 nm, were prepared by the method of Enoch & Strittmatter [Enoch, H., & Strittmatter, P. (1979) Proc. Natl. Acad. Sci. U.S.A. 76,...


    pub_type: 杂志文章


    authors: Teissie J,Tsong TY

    更新日期:1981-03-17 00:00:00

  • Basic and acidic regions flanking the HMG-box domain of maize HMGB1 and HMGB5 modulate the stimulatory effect on the DNA binding of transcription factor Dof2.

    abstract::The chromatin-associated high-mobility group (HMG) proteins of the plant HMGB family are characterized by a central HMG-box domain that is flanked by a basic N-terminal and an acidic C-terminal domain. By functional interaction with certain transcription factors, HMGB proteins contribute to transcriptional regulation....


    pub_type: 杂志文章


    authors: Grasser M,Christensen JM,Peterhänsel C,Grasser KD

    更新日期:2007-05-29 00:00:00

  • Active site specific cadmium(II)-substituted horse liver alcohol dehydrogenase: crystal structures of the free enzyme, its binary complex with NADH, and the ternary complex with NADH and bound p-bromobenzyl alcohol.

    abstract::Three crystal structures have been determined of active site specific substituted Cd(II) horse liver alcohol dehydrogenase and its complexes. Intensities were collected for the free, orthorhombic enzyme to 2.4-A resolution and for a triclinic binary complex with NADH to 2.7-A resolution. A ternary complex was crystall...


    pub_type: 杂志文章


    authors: Schneider G,Cedergren-Zeppezauer E,Knight S,Eklund H,Zeppezauer M

    更新日期:1985-12-03 00:00:00

  • Cloning of synthetic deoxyribonucleic acid that codes for embryonic cardiac myosin light-chain polypeptide.

    abstract::Double-stranded complementary deoxyribonucleic acid (cDNA) transcribed in vitro from a partially pure myosin light-chain messenger ribonucleic acid (mRNA) of the chick embryonic heart was cloned in Escherichia coli strain chi 1776 by using the HindIII cleavage site in the plasmid pBR322. The insertion of essentially f...


    pub_type: 杂志文章


    authors: Arnold HH,Siddiqui MA

    更新日期:1979-12-11 00:00:00

  • New synthetic cluster ligands for galactose/N-acetylgalactosamine-specific lectin of mammalian liver.

    abstract::Synthetic ligands containing up to six residues of nonreducing terminal galactose were prepared. The synthesis involved coupling of carboxyl groups of N-benzyloxy-carbonylaspartic acid or of N-benzyloxycarbonyltyrosyl-gamma-glutamylglutamic acid to the omega-amino group of the aglycon of a glycoside that contained up ...


    pub_type: 杂志文章


    authors: Lee RT,Lin P,Lee YC

    更新日期:1984-08-28 00:00:00

  • Catalytic mechanism of enterococcal kanamycin kinase (APH(3')-IIIa): viscosity, thio, and solvent isotope effects support a Theorell-Chance mechanism.

    abstract::Bacterial resistance to the aminoglycoside antibiotics is manifested primarily through the production of enzymes which covalently modify these drugs. The Enterococci and Staphylococci produce an ATP-dependent kinase, APH(3')-IIIa, which phosphorylates such antibiotics as kanamycin, amikacin, and neomycin, and this enz...


    pub_type: 杂志文章


    authors: McKay GA,Wright GD

    更新日期:1996-07-02 00:00:00

  • Characterization of hemopexin and its interaction with heme by differential scanning calorimetry and circular dichroism.

    abstract::Hemopexin is a plasma glycoprotein that has two structural domains (I and II) and binds and transports heme particularly to liver cells. Differential scanning calorimetry (DSC) studies show that hemopexin is largely stabilized by heme, which binds exclusively to domain I. The melting temperature (Tm) of heme-hemopexin...


    pub_type: 杂志文章


    authors: Wu ML,Morgan WT

    更新日期:1993-07-20 00:00:00

  • Identification of N(G)-methylarginine residues in human heterogeneous RNP protein A1: Phe/Gly-Gly-Gly-Arg-Gly-Gly-Gly/Phe is a preferred recognition motif.

    abstract::Three sites of N(G),N(G)-arginine methylation have been located at residues 205, 217, and 224 in the glycine-rich, COOH-terminal one-third of the HeLa A1 heterogeneous ribonucleoprotein. Together with the previously determined dimethylated arginine at position 193 [Williams et al., (1985) Proc. Natl. Acad. Sci. U.S.A....


    pub_type: 杂志文章


    authors: Kim S,Merrill BM,Rajpurohit R,Kumar A,Stone KL,Papov VV,Schneiders JM,Szer W,Wilson SH,Paik WK,Williams KR

    更新日期:1997-04-29 00:00:00

  • The binding of ATP and Mg2+ to the calcium adenosinetriphosphatase of sarcoplasmic reticulum follows a random mechanism.

    abstract::The enzyme form of the calcium adenosinetriphosphatase of sarcoplasmic reticulum (CaATPase) that is stable in the presence of calcium, cE.Ca2, has a binding site for the catalytic Mg2+ ion with a dissociation constant of 0.94 +/- 0.15 mM at 25 degrees C, pH 7.0, and 100 mM KCl. This is approximately 10 times smaller t...


    pub_type: 杂志文章


    authors: Reinstein J,Jencks WP

    更新日期:1993-07-06 00:00:00

  • Turkey gizzard caldesmon: molecular weight determination and calmodulin binding studies.

    abstract::Sedimentation equilibrium and sedimentation velocity measurements demonstrate that turkey gizzard caldesmon is an elongated molecule of molecular mass 75 +/- 2 kDa. The frictional ratio (2.14) is consistent with a prolate ellipsoid of axial ratio 24, corresponding to an apparent length and width of 516 and 21.5 A, res...


    pub_type: 杂志文章


    authors: Malencik DA,Ausio J,Byles CE,Modrell B,Anderson SR

    更新日期:1989-10-03 00:00:00

  • Structure of the cytoplasmic loop between putative helices II and III of the mannitol permease of Escherichia coli: a tryptophan and 5-fluorotryptophan spectroscopy study.

    abstract::In this work, four single tryptophan (Trp) mutants of the dimeric mannitol transporter of Escherichia coli, EII(mtl), are characterized using Trp and 5-fluoroTrp (5-FTrp) fluorescence spectroscopy. The four positions, 97, 114, 126, and 133, are located in a region shown by recent studies to be involved in the mannitol...


    pub_type: 杂志文章


    authors: Vos EP,Bokhove M,Hesp BH,Broos J

    更新日期:2009-06-16 00:00:00

  • Structure of the Rhodobacter sphaeroides light-harvesting 1 beta subunit in detergent micelles.

    abstract::The light harvesting 1 antenna (LH1) complex from Rhodobacter sphaeroides funnels excitation energy to the photosynthetic reaction center. Our ultimate goal is to build up the structure of LH1 from structures of its individual subunits, much as the antenna can self-assemble from its components in membrane-mimicking de...


    pub_type: 杂志文章


    authors: Sorgen PL,Cahill SM,Krueger-Koplin RD,Krueger-Koplin ST,Schenck CC,Girvin ME

    更新日期:2002-01-08 00:00:00

  • Loss of DNA minor groove interactions by exonuclease-deficient Klenow polymerase inhibits O6-methylguanine and abasic site translesion synthesis.

    abstract::The importance of DNA polymerase-DNA minor groove interactions on translesion synthesis (TLS) was examined in vitro using variants of exonuclease-deficient Klenow polymerase and site-specifically modified DNA oligonucleotides. Polymerase variant R668A lacks primer strand interactions, while variant Q849A lacks templat...


    pub_type: 杂志文章


    authors: Gestl EE,Eckert KA

    更新日期:2005-05-10 00:00:00

  • Engineering a novel specificity in subtilisin BPN'.

    abstract::The specificity of subtilisin BPN' toward substrates with large hydrophobic P4 residues has been improved by single amino acid replacements at positions 104 and 107. Mutations were designed to (i) increase the size of the P4 binding pocket by replacing Ile107, which is at the bottom of the S4 pocket, by Val, Ala, and ...


    pub_type: 杂志文章


    authors: Rheinnecker M,Baker G,Eder J,Fersht AR

    更新日期:1993-02-09 00:00:00

  • Backbone dynamics of a model membrane protein: 13C NMR spectroscopy of alanine methyl groups in detergent-solubilized M13 coat protein.

    abstract::The filamentous coliphage M13 possesses multiple copies of a 50-residue coat protein which is inserted into the inner membrane of Escherichia coli during infection. 13C nuclear magnetic resonance (NMR) spectroscopy has been used to probe the structure and dynamics of M13 coat protein solubilized in detergent micelles....


    pub_type: 杂志文章


    authors: Henry GD,Weiner JH,Sykes BD

    更新日期:1986-02-11 00:00:00