Showing papers by "David Eisenberg published in 1993"

Unconventional medicine in the United States. Prevalence, costs, and patterns of use.

Abstract: Background Many people use unconventional therapies for health problems, but the extent of this use and the costs are not known. We conducted a national survey to determine the prevalence, costs, and patterns of use of unconventional therapies, such as acupuncture and chiropractic. Methods We limited the therapies studied to 16 commonly used interventions neither taught widely in U.S. medical schools nor generally available in U.S. hospitals. We completed telephone interviews with 1539 adults (response rate, 67 percent) in a national sample of adults 18 years of age or older in 1990. We asked respondents to report any serious or bothersome medical conditions and details of their use of conventional medical services; we then inquired about their use of unconventional therapy. Results One in three respondents (34 percent) reported using at least one unconventional therapy in the past year, and a third of these saw providers for unconventional therapy. The latter group had made an average of 19 visits to suc...

Crystal structure of a synthetic triple-stranded alpha-helical bundle

Abstract: The x-ray crystal structure of a peptide designed to form a double-stranded parallel coiled coil shows that it is actually a triple-stranded coiled coil formed by three alpha-helices. Unlike the designed parallel coiled coil, the helices run up-up-down. The structure is stabilized by a distinctive hydrophobic interface consisting of eight layers. As in the design, each alpha-helix in the coiled coil contributes one leucine side chain to each layer. The structure suggests that hydrophobic interactions are a dominant factor in the stabilization of coiled coils. The stoichiometry and geometry of coiled coils are primarily determined by side chain packing in the solvent-inaccessible interior, but electrostatic interactions also contribute.

The structure of granulocyte-colony-stimulating factor and its relationship to other growth factors

Abstract: We have determined the three-dimensional structure of recombinant human granulocyte-colony-stimulating factor by x-ray crystallography. Phases were initially obtained at 3.0-A resolution by multiple isomorphous replacement and were refined by solvent flattening and by averaging of the electron density of the three molecules in the asymmetric unit. The current R factor is 21.5% for all data between 6.0- and 2.2-A resolution. The structure is predominantly helical, with 104 of the 175 residues forming a four-alpha-helix bundle. The only other secondary structure is also helical. In the loop between the first two long helices a four-residue 3(10)-helix is immediately followed by a 6-residue alpha-helix. Three residues in the short connection between the second and third bundle helices form almost one turn of left-handed helix. The up-up-down-down connectivity with two long crossover connections has been reported previously for five other proteins, which like granulocyte-colony-stimulating factor are all signaling ligands: growth hormone, granulocyte/macrophage-colony-stimulating factor, interferon beta, interleukin 2, and interleukin 4. Structural similarity among these growth factors occurs despite the absence of similarity in their amino acid sequences. Conservation of this tertiary structure suggests that these different growth factors might all bind to their respective sequence-related receptors in an equivalent manner.

Defensins promote fusion and lysis of negatively charged membranes

Abstract: Defensins, a family of cationic peptides isolated from mammalian granulocytes and believed to permeabilize membranes, were tested for their ability to cause fusion and lysis of liposomes. Unlike alpha-helical peptides whose lytic effects have been extensively studied, the defensins consist primarily of beta-sheet. Defensins fuse and lyse negatively charged liposomes but display reduced activity with neutral liposomes. These and other experiments suggest that fusion and lysis is mediated primarily by electrostatic forces and to a lesser extent, by hydrophobic interactions. Circular dichroism and fluorescence spectroscopy of native defensins indicate that the amphiphilic beta-sheet structure is maintained throughout the fusion process. Taken together, these results support the idea that protein-mediated membrane fusion depends not only on hydrophobic and electrostatic forces but also on the spatial arrangement of the amino acid residues to form a three-dimensional amphiphilic structure, which promotes the efficient mixing of the lipids between membranes. A molecular model for membrane fusion by defensins is presented, which takes into account the contributions of electrostatic forces, hydrophobic interactions, and structural amphiphilicity.

Cognitive Behavioral Techniques for Hypertension: Are They Effective?

Abstract: Purpose: To assess by analysis of published controlled trials the efficacy of cognitive behavioral therapies (such as biofeedback, relaxation, meditation) for essential hypertension. Data Identific...

Three-dimensional profiles from residue-pair preferences: identification of sequences with beta/alpha-barrel fold.

Abstract: The three-dimensional profile method expresses the three-dimensional structure of a protein as a table, the profile, which represents the local environment of each residue. The score of an amino acid sequence, aligned with the three-dimensional profile, reflects its compatibility with the profiled structure. In the original implementation, each local environment was characterized by its polarity, the area buried of its side chain, and its secondary structure. Here we describe a modified three-dimensional profile algorithm that characterizes the local environment in terms of the statistical preferences of the profiled residue for neighbors of specific residue types, main-chain conformations, or secondary structure. Combined profiles of the original and the three new types were tested on beta/alpha-barrel protein structures. The method identified the following enzymes of unknown three-dimensional structure as probable beta/alpha-barrels, all of which catalyze reactions in the biosynthesis of aromatic amino acids: anthranilate phosphoribosyltransferase (trpD), glutamine amidotransferase (trpG), and phosphoribosylformimino-5-aminoimidazole carboxamide ribotide isomerase (hisA).

Feedback inhibition of fully unadenylylated glutamine synthetase from Salmonella typhimurium by glycine, alanine, and serine.

Abstract: Bacterial glutamine synthetase (GS; EC 6312) was previously shown to be inhibited by nine end products of glutamine metabolism Here we present four crystal structures of GS, complexed with the substrate Glu and with each of three feedback inhibitors The GS of the present study is from Salmonella typhimurium, with Mn2+ ions bound, and is fully unadenylylated From Fourier difference maps, we find that L-serine, L-alanine, and glycine bind at the site of the substrate L-glutamate In our model, these four amino acids bind with the atoms they share in common (the "main chain" +NH3-CH-COO-) in the same positions Thus on the basis of our x-ray work, glycine, alanine, and serine appear to inhibit GS-Mn by competing with the substrate glutamate for the active site

Formation of the active site of ribulose-1,5-bisphosphate carboxylase/oxygenase by a disorder-order transition from the unactivated to the activated form

Abstract: Ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) catalyzes the key first step in photosynthetic CO2 fixation, the reaction that incorporates CO2 into sugar. In this study, refined crystal structures of unactivated tobacco RuBisCO and activated RuBisCO from spinach and tobacco, in complex with the reaction-intermediate analog 2-carboxyarabinitol 1,5-bisphosphate (CABP), are compared. Both plant enzymes are hexadecameric complexes of eight large and eight small subunits with a total relative molecular mass of approximately 550,000. The comparison of activated and unactivated forms of RuBisCO provides insight into the dynamics of action of this enzyme. The catalytic site, which is open to the solvent in the unactivated enzyme, becomes shielded in the activated CABP complex. This shielding is accomplished by a 12-A movement of the active-site "loop 6" (residues 331-338) and a disorder-order transition of three loops near the active-site entrance, the N terminus, the C terminus, and a loop comprising residues 64-68. All these residues belong to the catalytic large subunit. Domain rotations of about 2 degrees are observed, also tightening the active-site cleft. These observations provide an explanation for the extremely tight binding (Kd < or = 10(-11) M) of the CABP molecule. A striking correlation exists between crystallographic temperature factors in the activated enzyme and the magnitude of the atomic movement upon activation.

Crystal structure of activated tobacco rubisco complexed with the reaction-intermediate analogue 2-carboxy-arabinitol 1,5-bisphosphate.

Abstract: The crystal structure of activated tobacco rubisco, complexed with the reaction-intermediate analogue 2-carboxy-arabinitol 1,5-bisphosphate (CABP) has been determined by molecular replacement, using the structure of activated spinach rubisco (Knight, S., Andersson, I., & Branden, C.-I., 1990, J. Mol. Biol. 215, 113-160) as a model. The R-factor after refinement is 21.0% for 57,855 reflections between 9.0 and 2.7 A resolution. The local fourfold axis of the rubisco hexadecamer coincides with a crystallographic twofold axis. The result is that the asymmetric unit of the crystals contains half of the L8S8 complex (molecular mass 280 kDa in the asymmetric unit). The activated form of tobacco rubisco is very similar to the activated form of spinach rubisco. The root mean square difference is 0.4 A for 587 equivalent C alpha atoms. Analysis of mutations between tobacco and spinach rubisco revealed that the vast majority of mutations concerned exposed residues. Only 7 buried residues were found to be mutated versus 54 residues at or near the surface of the protein. The crystal structure suggests that the Cys 247-Cys 247 and Cys 449-Cys 459 pairs are linked via disulfide bridges. This pattern of disulfide links differ from the pattern of disulfide links observed in crystals of unactivated tobacco rubisco (Curmi, P.M.G., et al., 1992, J. Biol. Chem. 267, 16980-16989) and is similar to the pattern observed for activated spinach tobacco.

A model for oxidative modification of glutamine synthetase, based on crystal structures of mutant H269N and the oxidized enzyme.

Abstract: Proteolytic degradation of glutamine synthetase (GS) in Escherichia coli is known to follow "marking" by oxidative modification. At an early stage of the degradative pathway, oxidation of His 269 and Arg 344 abolishes GS enzymatic activity. We propose a mechanism for the early stage of oxidative inactivation of GS on the basis of the crystal structure of H269N and tryptophan fluorescence spectra of H269N and H269NR344Q: (1) Oxidation of Arg 344, adjacent to the n2 metal ion site, decreases ATP binding. (2) Oxidation of His 269 to Asn destroys the n2 site, consistent with the function of His 269 as a ligand for the n2 metal. (3) Loss of Mn2+ at the n2 site destroys the integrity of the ATP binding site. (4) Destruction of the ATP site results in the observed low enzymatic activity of H269N and H269NR344Q. During later stages of oxidative modification, the n1 metal ion site is destroyed and the active site of the enzyme becomes flexible as suggested by X-ray data collected from an oxidized crystal of GS. Thus, studies of mutant and oxidized enzymes confirm that there are at least two stages of oxidative modification of GS. These studies suggest that the early modification occurs at the n2 metal ion site, eliminating enzyme activity, and the later modification occurs at the n1 metal ion site, relaxing the GS structure, perhaps enabling proteolytic degradation. These studies also illuminate the differing roles of the two bound metal ions: the tightly bound n1 ion enhances the stability of the catalytically active conformation, and the less tightly bound n2 ion participates in ATP binding.

Crystal structure of the unactivated ribulose 1, 5-bisphosphate carboxylase/oxygenase complexed with a transition state analog, 2-carboxy-D-arabinitol 1, 5-bisphosphate

Abstract: The crystal structure of unactivated ribulose 1,5-bisphosphate carboxylase/oxygenase from Nicotiana tabacum complexed with a transition state analog, 2-carboxy-D-arabinitol 1,5-bisphosphate, was determined to 2.7 A resolution by X-ray crystallography. The transition state analog binds at the active site in an extended conformation. As compared to the binding of the same analog in the activated enzyme, the analog binds in a reverse orientation. The active site Lys 201 is within hydrogen bonding distance of the carboxyl oxygen of the analog. Loop 6 (residues 330-339) remains open and flexible upon binding of the analog in the unactivated enzyme, in contrast to the closed and ordered loop 6 in the activated enzyme complex. The transition state analog is exposed to solvent due to the open conformation of loop 6.

Novel diphtheria toxin-based molecules

Abstract: The invention features a chimeric diphtheria toxin molecule wherein all or part of a complementarity determining region of an antibody is inserted into a loop region of the Diphtheria toxin receptor binding-domain.