scholarly journals A computergraphical method of describing the shapes of subunit interfaces of oligomers. Analysis of the quaternary structure of concanavalin A and of prealbumin

1982 ◽  
Vol 205 (2) ◽  
pp. 353-359 ◽  
Author(s):  
E J Milner-White
Keyword(s):  
Concanavalin A ◽  
Quaternary Structure ◽  
Beta Sheets ◽  
New Method

A new method is described for mapping the shape of the intersubunit contacts of oligomers with identical subunits or pairs of subunits. Plots are generated for the dimer-dimer interactions of concanavalin A and prealbumin, which are both tetramers. Both contacts lie at the junction between the faces of two apposed beta-sheets, but in most other respects the interactions are seen to be different.

Alteration of quaternary structure and biological activity of concanavalin A

1984 ◽  
Vol 3 (1) ◽  
pp. 63-71 ◽  
Author(s):  
Shin-ichi Ishii ◽  
Yukichi Abe ◽  
Isao Tanaka ◽  
Masaki Saito
Keyword(s):  
Biological Activity ◽  
Concanavalin A ◽  
Quaternary Structure

Concanavalin A-horseradish peroxidase bridge staining of alpha-1 glycoproteins separated by isoelectric focusing on polyacrylamide gel.

1976 ◽  
Vol 24 (8) ◽  
pp. 908-914 ◽  
Author(s):  
R C Allen ◽  
S S Spicer ◽  
D Zehr
Keyword(s):  
Horseradish Peroxidase ◽  
Isoelectric Focusing ◽  
Concanavalin A ◽  
Periodic Acid ◽  
Polyacrylamide Gel ◽  
New Method ◽  
Polyacrylamide Gels ◽  
Periodic Acid Schiff ◽  
Coomassie Blue

The Coomassie Blue protein stain and the periodic acid-Schiff stain for glycoproteins are compared to a new method of staining glycoproteins resolved electrophoretically. The method utilizes a Concanavalin A-horseradish peroxidase sequence to visualize selectively glycoproteins with terminal or internal mannose or terminal N-acetylglucosamine. The method applied to characterization of M and Z allele products of alpha-l-antitrypsins separated by isoelectric focusing of polyacrylamide gels slabs have revealed differences in carbohydrate content of various components that were previously undetected.

Differential ultracentrifugation coupled to small-angle X-ray scattering on macromolecular complexes

2015 ◽  
Vol 48 (3) ◽  
pp. 769-775 ◽  
Author(s):  
Robert M. G. Hynson ◽  
Anthony P. Duff ◽  
Nigel Kirby ◽  
Stephan Mudie ◽  
Lawrence K. Lee
Keyword(s):  
Small Angle ◽  
Quaternary Structure ◽  
Protein Complexes ◽  
New Method ◽  
Size Exclusion ◽  
Saxs Data ◽  
X Ray ◽  
X Ray Scattering ◽  
Macromolecular Protein ◽  
Ray Scattering

Small-angle X-ray scattering (SAXS) can provide accurate structural information and low-resolution shapes of macromolecules in solution. The technique is particularly amenable to large protein assemblies, which produce a strong scattering signal. Hence, SAXS can be a powerful tool to elucidate quaternary structure, especially when used in combination with high-resolution structural techniques such as X-ray crystallography and NMR. Sample requirements for SAXS experiments are stringent and only monodispersed samples can be satisfactorily analysed. Often, it is not possible to obtain a stable monodispersed sample of the protein of interest, in particular for multi-subunit protein complexes. In these circumstances, when the complex is less than approximately 1 MDa, size exclusion chromatography (SEC) coupled with SAXS (SEC-SAXS) can facilitate the separation of monodispersed protein from a polydispersed sample for a sufficient amount of time to collect useful SAXS data. However, many very large multi-subunit macromolecular assemblies have not been successfully purified with SEC, and hence despite being well suited to SAXS there is often no way to produce sample of sufficient quality. Rather than SEC, differential ultracentrifugation (DU) is the method of choice for the final step in the purification of large macromolecular protein complexes. Here, a new method is described for collecting SAXS data on samples directly from the fractionated elution of ultracentrifuge tubes after DU. It is demonstrated using apoferritin as a model protein that, like SEC-SAXS, DU-coupled SAXS can facilitate simultaneous purification and data collection. It is envisaged that this new method will enable high-quality SAXS data to be collected on a host of large macromolecular protein complex assemblies for the first time.

scholarly journals Automated evaluation of quaternary structures from protein crystals

2017 ◽  
Author(s):  
Spencer Bliven ◽  
Aleix Lafita ◽  
Althea Parker ◽  
Guido Capitani ◽  
Jose M Duarte
Keyword(s):  
Protein Data Bank ◽  
Quaternary Structure ◽  
Protein Structures ◽  
Data Bank ◽  
New Method ◽  
Protein Assemblies ◽  
Native Solution ◽  
Biologically Relevant ◽  
Crystal Contacts ◽  
Structure Of Proteins

AbstractA correct assessment of the quaternary structure of proteins is a fundamental prerequisite to understanding their function, physico-chemical properties and mode of interaction with other proteins. Currently about 90% of structures in the Protein Data Bank are crystal structures, in which the correct quaternary structure is embedded in the crystal lattice among a number of crystal contacts. Computational methods are required to 1) classify all protein-protein contacts in crystal lattices as biologically relevant or crystal contacts and 2) provide an assessment of how the biologically relevant interfaces combine into a biological assembly In our previous work we addressed the first problem with our EPPIC (Evolutionary Protein Protein Interface Classifier) method. Here, we present our solution to the second problem with a new method that combines the interface classification results with symmetry and topology considerations. The new algorithm enumerates all possible valid assemblies within the crystal using a graph representation of the lattice and predicts the most probable biological unit based on the pairwise interface scoring. Our method achieves 85% precision on a new dataset of 1,481 biological assemblies with consensus of PDB annotations. Although almost the same precision is achieved by PISA, currently the most popular quaternary structure assignment method, we show that, due to the fundamentally different approach to the problem, the two methods are complementary and could be combined to improve biological assembly assignments. The software for the automatic assessment of protein assemblies (EPPIC version 3) has been made available through a web server at http://www.eppic-web.org.Author summaryX-ray diffraction experiments are the main experimental technique to reveal the detailed atomic 3-dimensional structure of proteins. In these experiments, proteins are packed into crystals, an environment that is far away from their native solution environment. Determining which parts of the structure reflect the protein’s state in the cell rather than being artifacts of the crystal environment can be a difficult task. How the different protein subunits assemble together in solution is known as the quaternary structure. Finding the correct quaternary structure is important both to understand protein oligomerization and for the understanding of protein-protein interactions at large. Here we present a new method to automatically determine the quaternary structure of proteins given their crystal structure. We provide a theoretical basis for properties that correct protein assemblies should possess, and provide a systematic evaluation of all possible assemblies according to these properties. The method provides a guidance to the experimental structural biologist as well as to structural bioinformaticians analyzing protein structures in bulk. Assemblies are provided for all proteins in the Protein Data Bank through a public website and database that is updated weekly as new structures are released.

A new method for the partial purification of leishmania amastigotes from cutaneous lesions

Parasitology ◽  
1980 ◽  
Vol 80 (1) ◽  
pp. 105-112 ◽  
Author(s):  
R. B. Infante ◽  
A. G. Hernandez ◽  
F. Riggione ◽  
K. Dawidowicz
Keyword(s):  
Dna Replication ◽  
Cell Count ◽  
Concanavalin A ◽  
Growth Rates ◽  
New Method ◽  
Partial Purification ◽  
Leishmania Braziliensis ◽  
Cutaneous Lesions ◽  
Apparent Loss ◽  
Leishmania Amastigotes

SummaryAmastigotes from Leishmania braziliensis and L. mexicana were isolated from cutaneous lesions in infected animals using the plant lectin Concanavalin A as a specific agglutination agent. Amastigotes were collected in preparations of up to 95% purity as determined by cell count. The parasites obtained by this method showed no apparent loss of viability as measured by growth rates and DNA replication, or pathogenicity as measured by routine passages in hamsters or mice.

scholarly journals Freeze-fracture cytochemistry: thin sections of cells and tissues after labeling of fractures faces.

1981 ◽  
Vol 29 (8) ◽  
pp. 917-928 ◽  
Author(s):  
P P da Silva ◽  
C Parkison ◽  
N Dwyer
Keyword(s):  
Concanavalin A ◽  
Binding Sites ◽  
Freeze Fracture ◽  
New Method ◽  
Isolated Cells ◽  
Colloidal Iron ◽  
Thin Sections ◽  
Combined Application ◽  
Section Electron ◽  
Anionic Binding Sites

Experimental details of a new method for the cytochemical characterization of the membrane faces and cytoplasm produced by freeze-fracture of isolated cells and tissues are presented. This new method-"fracture-label"-involves grinding of frozen samples immersed in liquid nitrogen, thawing, cytochemical labeling of the fractured faces, and processing for thin section electron microscopy. Cationized ferritin (at pH. 7.5 and 4.0), colloidal iron, as well as concanavalin A are used to label the fractures faces of leukocytes and Hela cells embedded in a cross-linked matrix of bovine serum albumin and of liver and spleen tissues. Our results show the presence of numerous anionic binding sites on the fracture faces of all plasma and cytoplasmic membranes, and of concanavalin A binding sites preferentially associated to the exoplasmic fracture faces of plasma and nuclear envelope membranes. A proportion of the anionic sites appears to be revealed by, or during, the freeze-fracture process. Colloidal iron labeling also shows preferential association with the chromatin areas of cross-fractured nuclei. The results show that "fracture-label", i.e., the combined application of freeze-fracture and cytochemical labeling techniques, can be used to study the surface chemistry of the fractures faces of biological membranes as well as of cross-fractured cytoplasm.

Quaternary structure of Limulus polyphemus hemocyanin

1978 ◽  
Vol 36 (2) ◽  
pp. 176-177
Author(s):  
T. Wichertjes ◽  
E.J. Kwak ◽  
E.F.J. Van Bruggen
Keyword(s):  
Electron Microscopy ◽  
Functional Properties ◽  
Horseshoe Crab ◽  
Temperature Jump ◽  
Quaternary Structure ◽  
Crystallographic Analysis ◽  
Limulus Polyphemus ◽  
Oxygen Binding ◽  
X Ray ◽  
Intact Molecule

Hemocyanin of the horseshoe crab (Limulus polyphemus) has been studied in nany ways. Recently the structure, dissociation and reassembly was studied using electron microscopy of negatively stained specimens as the method of investigation. Crystallization of the protein proved to be possible and X-ray crystallographic analysis was started. Also fluorescence properties of the hemocyanin after dialysis against Tris-glycine buffer + 0.01 M EDTA pH 8.9 (so called “stripped” hemocyanin) and its fractions II and V were studied, as well as functional properties of the fractions by NMR. Finally the temperature-jump method was used for assaying the oxygen binding of the dissociating molecule and of preparations of isolated subunits. Nevertheless very little is known about the structure of the intact molecule. Schutter et al. suggested that the molecule possibly consists of two halves, combined in a staggered way, the halves themselves consisting of four subunits arranged in a square.

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