ISSN:0887-3585    

DOI:10.1002/prot.340060202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractThe results of a 125 psec molecular dynamics simulation of alacheadpiece–operator complex in aqueous solution are reported. The complexsatisfies essentially all experimental distance information derived from two‐dimensional nuclear magnetic resonance (2‐D‐NMR) studies. The interaction betweenlacrepressor headpiece and its operator based on many direct‐ and water‐mediated hydrogenbonds and nonpolar contacts which allow the formation of a tight complex. Nostable hydrogen bonds between side chains and bases and found, while specific contacts occur between both nonpolar groups and, to a lesserextent, through water‐mediated hydrogen bonds. The simulated complex structure in water is intrinsically stable without application of nuclear Overhauser effect (NOE) distance restraints, while being compatible with most of the available biochemical, genetic, andchemically induced dynamic nuclear polarization

ISSN:0887-3585    

DOI:10.1002/prot.340060203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractPrevious structure/function analyses of the DNA repairenzyme, T4 endonuclease V, have suggested that the extreme carboxyl portion of the enzyme is associated with pyrimidine dimer‐specific binding (Recinos and Lloyd, and Stump and Lloyd, Biochemistry 27:1832–1838 and 1839–1843, 1988, respectively). Within the final 11 amino acids there are 5 aromatic, 2 basic, and no acidic residues and it has been proposed that these residues stack with and electrostatically interact with the kinked DNA at the site of a pyrimidine dimer. The role of the tyrosine residue at position 129 has beeninvestigated by oligonucleotide site‐directed mutagenesis in which the codon for Tyr‐129 has been altered to reflect conservative changesof Trp and Phe and more dramatic changes of Ser‐ a stop, codon, deletion of the codon or introduction of a frameshift. Both changesto the aromatic amino acids resulted in proteins which accumulated will inE. coliand not only significantly enhanced the UV survival of repair, deficient cells but also complemented a defectivedenV gene within UV‐irradiated T4 phage. Partially purified preparations of the Tyr‐129 → Trp and Tyr‐129 → Phe mutants were assayed for their ability to processively incise UV‐irradiated plasmid DNA (a nicking reaction carried out at low 25 mM salt concentrations). The mutant enzymes Tyr‐129 → Phe and Tyr‐129 → Trp displayed a 1000% and 500% enhanced specific nicking activity, respectively. These reactions were also shown to be completely processive. Assays performed at higher (100 mM) salt concentrations reduced the specific activities of the mutant enzymes approximately to that of wild type for the Tyr‐129 → Phe mutant and to 20% that of wil

ISSN:0887-3585    

DOI:10.1002/prot.340060204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractA recombinant plasmid encompassing the human immunodeficiency virus type 1 (HIV 1) protease coding sequence and flanking regions (Ala‐13 to Gly‐185 of thepolopen reading frame) has been expressed in two distinct strains ofEscherichia coli, AR58 and AR68. In the first strain, AR58, the primary translation product, a 25 kilodalton (kDa) precursor protein, is short‐lived and rapidly processes itself to the 11 kDa mature protease in vivo. In the second strain, AR68, the 25 kDa species isonly partially processed, and it, a 13 kDA intermediate, and the mature 11 kDA enzyme accumulate at a ratio of 3:4.5:2.5, respectively. The 11 kDa mature protease from AR58 and the 25 kDa precursor from AR68 have been purified to homogeneity. The yield of 11 kDa enzyme from AR58 is approximately 0.02 mg/g wet weight ofE. colicell pellet. The protease has both the expected NH2‐ and COOH‐terminal sequences. The yield of 25 kDa enzyme from AR68 is approximately 0.1 mg/g wet weight ofE. colicell pellet. In vitro, the 25 kDa precursor enzyme rapidly (t1/2≅ 9 min) processes itself into a species with a mass of ∼13kDa and a species with a mass of ∼11 kDa. Both of these latter species can be separated by RP‐HPLC, have the NH2‐terminal sequence expected for the mature protease, and are active. The 11 kDa enzyme from AR58 comigrates with the 11 kDa enzyme from AR68 on RP‐HPLC and SDS poly acrylamide gel electrophoresis. On extended incubation at 4°C at either neutral or acidic pH all species of the proteinexhibit further autodegradation at defined sequences. The availability of the mature, 11 kDa enzyme and the 25 kDa precursor will allow biochemical and physical studies on this

ISSN:0887-3585    

DOI:10.1002/prot.340060205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractAn efficient algorithm was characterized that determines the similarity in main chain conformation between short protein substructures. The algorithm computes Δt, the root mean square difference in ϕ and ψ torsion angles over a small number of amino acids (typically 3–5). Using this algorithm, large number of protein substrates comparisons were feasible. The parameter Δtwas sensitive to variations in local protein conformation, and it correlates with Δr, the root mean square deviation in atomic coordinates. Values for Δtwere obtained that define similarity thresholds, which determine whether two substructure are considered structurally similar. To set a lower bound on the similarity threshold, we estimated the component of Δtdue to measurement noise fromcomparisons of independently refined coordinates of the same protein. A sample distribution of Δtfrom nonhomologous protein comparisons identified an upper bound on the similarity threshold, one that refrains from incorporating large numbers of nonmatching comparisons large numbers of nonmatching comparisons. Unlike methods based on Cα atoms alone, Δtwas sensitive to rotations in the peptide plane, shown to occur in several proteins. Comparisons of homologus proteins by Δtshowed that the active site torsion angles are highly conserved. The Δtmethod was applied to the α‐chain of human hemoglobin, where it readily demonstrated the local differences in the structures of different

ISSN:0887-3585    

DOI:10.1002/prot.340060206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractEquilibrium geometries and binding energies of model “salt” or “ion” bridge systems have been computed by ab initio quantum chemistry techniques (GAUSSIAN82) and by empirical techniques (AMBER2.0). Formate and dimethyl phosphate served as anions in the model compounds while interacting with several organic cations, including methyl ammonium, methyl guanidinium, and divalent metal ion (either Mg2+or Ca2+) without and with an additional chloride; and a divalent metal ion (either Mg2+or Ca2+), chloride, and four water molecules of hydration about the metal ion. The majority of the quantum chemical computations were performed using a split‐valence basis set. For the model compounds studied we find that the ab initio geometries are in remarkably goodagreement with the molecular mechanics geometries.Several Calculations werealso performed using diffuse fractions. The formate anion binds these modelcations more strongly than does dimethyl phosphate, while the organiccation methyl ammonium binds model anions more strongly than does methyl guanidinium. Finally, in model compounds including organic anions, Mg2+or Ca2+and four molecules of water, and a chloride anion, we find that the equilibrium structure of the magnesium complex involves a solvent separated ion pair (the magnesium ion is six coordinate), whereas the calcium ion complex remains seven coordinate. Molecular mechanics overestimates binding energies, but the estimates may be close enough to actual binding energies togive useful insight into the details energies to give useful insight into the details of salt bridges in biologica

ISSN:0887-3585    

DOI:10.1002/prot.340060207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

摘要:

AbstractThe current work describes a simplified representation of protein structure with uses in the simulation of protein folding. The model assumes that a protein can be represented by a freely rotating rigid chain with a single atom approximately the effect of each side chains. Potentials describing the attraction or repulsion between different types of amino acids are determined directly from the distribution of amino acids in the database of known protein structures. The optimization technique of simulated annealinghas been used to dynamically sample the conformations available to this sample model, allowing the protein to evolve from an extended, random coil into a compact globular structure. Many characteristics expected of true proteins, such as the sequence‐dependent formation of secondary structure, the partitioning of hydrophobic residues, and specific disulfide, suggestion the model may accurately simulate the folding proces

ISSN:0887-3585    

DOI:10.1002/prot.340060208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY

ISSN:0887-3585    

DOI:10.1002/prot.340060201

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1989

数据来源: WILEY