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Journal Issue - Volume 4 Issue 11 (November 1995)

Abstract An important, but often neglected, contribution to the thermodynamics of protein folding is the loss of entropy that results from restricting the number of accessible side‐chain conformers in the native structure. Conformational entropy changes can be found by comparing the number of accessible rotamers in the unfolded and folded states, or by estimating fusion entropies. Comparison of several sets of results using...

Abstract The Alacoil is an antiparallel (rather than the usual parallel) coiled‐coil of α‐helices with Ala or another small residue in every seventh position, allowing a very close spacing of the helices (7.5–8.5 Å between local helix axes), often over four or five helical turns. It occurs in two distinct types that differ by which position of the heptad repeat is occupied by Ala and by whether the closest points on the backbone of...

Abstract The consequences of site‐directed mutagenesis experiments are often anticipated by empirical rules regarding the expected effects of a given amino acid substitution. Here, we examine the effects of “conservative” and “nonconservative” substitutions on the X‐ray crystal structures of human recombinant FKBP12 mutants in complex with the immunosuppressant drug FK506 (tacrolimus). R42K and R42I mutant complexes show 110‐fold...

Abstract Homeodomains are 60 amino acid DNA binding domains found in numerous eukaryotic transcription factors. The homeodomain family is a useful system for studying sequence‐structure relationships because several hundred sequences are known and the structures of several homeodomains have been determined. Covariation of amino acid residues in the homeodomain family has been investigated to see whether strongly covariant residue...

Abstract We have converted the Drosophila engrailed homeodomain into a sequence‐specific nuclease by linking the protein to the chemical nuclease 1,10‐phenanthroline‐copper (OP‐Cu). Unique cysteines were introduced at six positions into the homeodomain by site‐directed mutagenesis for the covalent attachment of OP‐Cu. The varied DNA‐binding affinity and specificity of these mutants and the DNA cleavage pattern of their OP‐Cu...

Abstract Network‐editing experiments are variants of the basic NOESY experiment that allow more accurate direct measurement of interproton distances in macromolecules by defeating specific spin‐diffusion pathways. Two network‐editing approaches, block‐decoupled NOESY and complementary‐block‐decoupled‐NOESY, were applied as three‐dimensional, heteronuclear‐edited experiments to distance measurement in a small protein, turkey...

Abstract In recent years, it has been repeatedly demonstrated that the coordinates of the main‐chain atoms alone are sufficient to determine the side‐chain conformations of buried residues of compact proteins. Given a perfect backbone, the side‐chain packing method can predict the side‐chain conformations to an accuracy as high as 1.2 Å RMS deviation (RMSD) with greater than 80% of the χ angles correct. However, similarly rigorous...

Abstract Six single‐Trp mutants were engineered by individually reintroducing each of the native Trp residues into a functional lactose permease mutant devoid of Trp (Trp‐less permease; Menezes ME, Roepe PD, Kaback HR, 1990, Proc Natl Acad Sci USA 87:1638–1642), and fluorescent properties were studied with respect to solvent accessibility, as well as alterations produced by ligand binding. The emission of Trp 33, Trp 78, Trp 171, and Trp 233 is...

Abstract A few studies indirectly support the existence of an intermediate in the transition of Ca2+‐saturated bovine α‐lactalbumin (α‐LA) from the native (N) to the acidic (A) state, known as the molten globule state. However, direct experimental evidence for the appearance of this intermediate has not been obtained. The signal of circular polarization of luminescence (CPL) is sensitive to fine conformational transitions because of its...

Abstract The Escherichia coli periplasmic dipeptide binding protein functions in both peptide transport and taxis toward peptides. The structure of the dipeptide binding protein in complex with Gly‐Leu (glycyl‐L‐leucine) has been determined at 3.2 Å resolution. The binding site for dipeptides is designed to recognize the ligand's backbone while providing space to accommodate a variety of side chains. Some repositioning of protein...

Abstract The structure and function of the periplasmic heme‐binding protein HbpA of Haemophilus influenzae were investigated. This protein is involved in the import of heme into the bacteria through the inner membrane, and thus is a key element of the organism's ability to survive in blood. A high degree of sequence similarity between HbpA and the dipeptide‐binding protein of Escherichia coli is suggested to be the result of a functional...

Abstract The crystallographic structure of bovine prothrombin fragment 1 bound with calcium ions was used to construct the corresponding human prothrombin structure (hf1/Ca). The model structure was refined by molecular dynamics to estimate the average solution structure. Accommodation of long‐range ionic forces was essential to reach a stable solution structure. The γ‐carboxyglutamic acid (Gla) domain and the kringle domain of...

Abstract Thermal denaturation of Torpedo californica acetylcholinesterase, a disulfide‐linked homodimer with 537 amino acids in each subunit, was studied by differential scanning calorimetry. It displays a single calorimetric peak that is completely irreversible, the shape and temperature maximum depending on the scan rate. Thus, thermal denaturation of acetylcholinesterase is an irreversible process, under kinetic control, which is...

Abstract Glutamine synthetase (GS) catalyzes the ATP‐dependent condensation of ammonia and glutamate to yield glutamine, ADP, and inorganic phosphate in the presence of divalent cations. Bacterial GS is an enzyme of 12 identical subunits, arranged in two rings of 6, with the active site between each pair of subunits in a ring. In earlier work, we have reported the locations within the funnel‐shaped active site of the substrates...

Abstract A homology model for the pig isozyme of the pyridoxal phosphate‐dependent enzyme γ‐aminobutyrate (GABA) aminotransferase has been built based mainly on the structure of dialkylglycine decarboxylase and on a multiple sequence alignment of 28 evolutionarily related enzymes. The proposed active site structure is presented and analyzed. Hypothetical structures for external aldimine intermediates explain several characteristics...

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