Search results for "protein conformation"
showing 10 items of 515 documents
Direct identification of the agonist binding site in the human brain cholecystokinin-B receptor
1999
In investigating the agonist binding site of the human brain cholecystokininB receptor (CCKBR), we employed the direct protein chemical approach using a photoreactive tritiated analogue of sulfated cholecystokinin octapeptide, which contains the p-benzoylbenzoyl moiety at the N-terminus, followed by purification of the affinity-labeled receptor to homogeneity. This probe bound specifically, saturably, and with high affinity (KD = 1.2 nM) to the CCKBR and has full agonistic activity. As the starting material for receptor purification, we used stably transfected HEK 293 cells overexpressing functional CCKBR. Covalent labeling of the WGA-lectin-enriched receptor revealed a 70-80 kDa glycoprote…
Activation of bee venom phospholipase A2 through a peptide-enzyme complex
1995
AbstractPhospholipase A2 activation by membrane-bound peptides was investigated in order to understand the role of the membrane-induced conformation on activation, and to examine the occurrence of a peptide-enzyme complex at the lipid/water interface. For the peptides studies, bee venom phospholipase A2 was stimulated regardless of the membrane-bound conformation (α-helix, β-sheet or random coil). Using antisera raised against melittin, we were able to demonstrate the occurrence of a calcium-dependent complex involving the enzyme, phospholipid substrate, and peptide.
Hemoglobin dynamics in rat erythrocytes investigated by M�ssbauer spectroscopy
1991
Rats have been enriched in 57Fe and erythrocytes were isolated from the blood. Mössbauer absorption spectroscopy on the hemoglobin of these erythrocytes has shown rather similar dynamics as found earlier in crystals of myoglobin, in frozen solutions of human hemoglobin and in a number of other proteins. The results strongly indicate that the motion of the heme and presumably some part of the F-helix is mainly influenced by the average viscosity of the sample determined by a network of hydrogen bridges and other weak interactions. Extrapolations of Mössbauer results from protein crystals to proteins in their physiological surroundings seem to be suitable for heme proteins.
Expression of Active Streptolysin O in Escherichia coli as a Maltose-Binding-Protein-Streptolysin-O Fusion Protein. The N-Terminal 70 Amino Acids are…
1996
Streptolysin 0 (SLO) is the prototype of a family of cytolysins that consists of proteins which bind to cholesterol and form very large transmembrane pores. Structure/function studies on the pore-forming cytolysin SLO have been complicated by the proteolytic inactivation of a substantial portion of recombinant SLO (rSLO) expressed in Escherichia coli. To overcome this problem, translational fusions between the E. coli maltose-binding protein (MBP) gene and SLO were constructed, using the vectors pMAL-p2 and pMAL-c2. MBP-SLO fusion proteins were degraded if secreted into the E. coli periplasm, but intact, soluble MBP-SLO fusion proteins were produced at high levels in the cytoplasm. Active S…
Analysis of Vibrational Circular Dichroism Spectra of Peptides
2020
Vibrational circular dichroism (VCD) is one of the major spectroscopic tools to study peptides. Nevertheless, a full understanding of what determines the signs and intensities of VCD bands of these compounds in the amide I and amide II spectral regions is still far from complete. In the present work, we study the origin of these VCD signals using the general coupled oscillator (GCO) analysis, a novel approach that has recently been developed. We apply this approach to the ForValNHMe model peptide in both α-helix and β-sheet configurations. We show that the intense VCD signals observed in the amide I and amide II spectral regions essentially have the same underlying mechanism, namely, the th…
Probing ensemble polymorphism and single aggregate structural heterogeneity in insulin amyloid self-assembly.
2020
Ensembles of protein aggregates are characterized by a nano- and micro-scale heterogeneity of the species. This diversity translates into a variety of effects that protein aggregates may have in biological systems, both in connection to neurodegenerative diseases and immunogenic risk of protein drug products. Moreover, this naturally occurring variety offers unique opportunities in the field of protein-based biomaterials. In the above-mentioned fields, the isolation and structural analysis of the different amyloid types within the same ensemble remain a priority, still representing a significant experimental challenge. Here we address such complexity in the case of insulin for its relevance…
Glutathione peroxidase-1 in health and disease: from molecular mechanisms to therapeutic opportunities.
2011
Reactive oxygen species, such as superoxide and hydrogen peroxide, are generated in all cells by mitochondrial and enzymatic sources. Left unchecked, these reactive species can cause oxidative damage to DNA, proteins, and membrane lipids. Glutathione peroxidase-1 (GPx-1) is an intracellular antioxidant enzyme that enzymatically reduces hydrogen peroxide to water to limit its harmful effects. Certain reactive oxygen species, such as hydrogen peroxide, are also essential for growth factor-mediated signal transduction, mitochondrial function, and maintenance of normal thiol redox-balance. Thus, by limiting hydrogen peroxide accumulation, GPx-1 also modulates these processes. This review explor…
Crystal Structure of the Maturation Protein from Bacteriophage Qβ
2017
Abstract Virions of the single-stranded RNA bacteriophages contain a single copy of the maturation protein, which is bound to the phage genome and is required for the infectivity of the particles. The maturation protein mediates the adsorption of the virion to bacterial pili and the subsequent release and penetration of the genome into the host cell. Here, we report a crystal structure of the maturation protein from bacteriophage Qβ. The protein has a bent, highly asymmetric shape and spans 110 A in length. Apart from small local substructures, the overall fold of the maturation protein does not resemble that of other known proteins. The protein is organized in two distinct regions, an α-he…
Development and characterization of a 293 cell line with regulatable expression of the hepatitis B virus large envelope protein
2004
During the life cycle of hepatitis B virus (HBV) the large L envelope protein plays a pivotal role that is related to its peculiar dual transmembrane topology. To study the complex structure and diverse functions of L under regulated conditions of production, a human 293 cell line stably expressing L under the control of the ecdysone-inducible promoter was generated. Cells demonstrated stringent dose- and time-dependent kinetics of induction with undetectable background expression in the absence of the inducer. Temporal control of L expression allowed to trace (i) its posttranslational reorientation resulting in the mixed topology; (ii) its spatial redistribution from the endoplasmic reticu…
The diversity of GABAA receptors. Pharmacological and electrophysiological properties of GABAA channel subtypes.
1998
The amino acid gamma-aminobutyric-acid (GABA) prevails in the CNS as an inhibitory neurotransmitter that mediates most of its effects through fast GABA-gated Cl(-)-channels (GABAAR). Molecular biology uncovered the complex subunit architecture of this receptor channel, in which a pentameric assembly derived from five of at least 17 mammalian subunits, grouped in the six classes alpha, beta, gamma, delta, sigma and epsilon, permits a vast number of putative receptor isoforms. The subunit composition of a particular receptor determines the specific effects of allosterical modulators of the GABAARs like benzodiazepines (BZs), barbiturates, steroids, some convulsants, polyvalent cations, and et…