scholarly journals An examination of the utility of photogenerated reagents by using α-chymotrypsin

1974 ◽  
Vol 143 (3) ◽  
pp. 663-668 ◽  
Author(s):  
Alex J. Bridges ◽  
Jeremy R. Knowles
Keyword(s):  
Tertiary Structure ◽  
Protein Molecule ◽  
Aryl Azide ◽  
Tryptic Fragment ◽  
High Degree ◽  
Nitrophenyl Ester ◽  
Labelling Method ◽  
Binding Locus ◽  
Covalently Bound

As a test of the labelling characteristics of photogenerated reagents, an aryl azide was photolysed in the aromatic-binding locus of a protein of known tertiary structure. The acyl-enzyme derived from the reaction of α-chymotrypsin with the p-nitrophenyl ester of p-azido[14C]cinnamate was isolated and photolysed. About 60% of the acyl group is covalently bound to the protein after photolysis and deacylation, and labelled enzyme is inactive. The covalently attached label is localized in the C chain of chymotrypsin, and there are firm indications that the major labelled tryptic fragment of the C chain is that which constitutes the aromatic-binding locus of the enzyme. The high degree of labelling of that portion of the protein molecule predicted on the basis of the known chemistry and structure of α-chymotrypsin, provides gratifying confirmation of the utility of the photo-labelling method.

Subunit δ of Chloroplast F0F1 ATPase and OSCP of Mitochondrial F0F1 ATPase: a Comparison by CD-Spectroscopy

1991 ◽  
Vol 46 (9-10) ◽  
pp. 759-764 ◽  
Author(s):  
Siegfried Engelbrecht ◽  
Jennifer Reed ◽  
François Penin ◽  
Danièle C. Gautheron ◽  
Wolfgang Junge
Keyword(s):  
Primary Structure ◽  
Tertiary Structure ◽  
Atp Synthesis ◽  
Structural Similarity ◽  
Cd Spectroscopy ◽  
Α Helix ◽  
And Function ◽  
High Degree ◽  
Very High ◽  
Secondary And Tertiary Structure

Abstract CD spectra have been recorded with subunit δ from chloroplast CF0CF1 and with OSCP from mitochondrial MF0MF1. These subunits are supposed to act similarly at the interface between proton transport through the F0-portion and ATP-synthesis in the F1-portion of their respective F0F1-ATPase. Evaluation of the data for both proteins revealed a very high α-helix content of -85% and practically no β-sheets. Despite their low homology on the primary structure level (23% identity) and their different electrostatic properties (pl-values differ by 3 units), spinach δ and porcine OSCP are indistinguishable with respect to their secondary structure as measured by CD. Prediction and analysis of consensual a-helices even in poorly conserved regions indicate α high degree of structural similarity between chloroplast δ and OSCP. In view of the topology and function of δ and OSCP in intact F0F1 these findings are interpreted to indicate the dominance of secondary and tertiary structure over the primary structure in their supposed function between proton flow and ATP-synthesis.

Fractal Analysis of Tertiary Structure of Protein Molecule

1984 ◽  
Vol 53 (6) ◽  
pp. 2162-2171 ◽  
Author(s):  
Yoshinori Isogai ◽  
Toshiyuki Itoh
Keyword(s):  
Fractal Analysis ◽  
Tertiary Structure ◽  
Protein Molecule

scholarly journals Preparation of uniformly labelled 13C- and 15N-plants using customised growth chambers

2020 ◽  
Author(s):  
Asja Ceranic ◽  
Maria Doppler ◽  
Christoph Büschl ◽  
Alexandra Parich ◽  
Kangkang Xu ◽  
...  
Keyword(s):  
Durum Wheat ◽  
Plant Material ◽  
Life Science ◽  
Plant Stress ◽  
Science Research ◽  
Nutrient Supply ◽  
Technical Equipment ◽  
Growth Chambers ◽  
High Degree ◽  
Labelling Method

Abstract Background Stable isotopically labelled organisms found wide application in life science research including plant physiology, plant stress and defense as well as metabolism related sciences. Therefore, the reproducible production of plant material enriched with stable isotopes such as 13 C and 15 N is of considerable interest. A high degree of enrichment (>96 atom%) with a uniformly distributed isotope (global labelling) is accomplished by a continuous substrate supply during plant growth/cultivation. In the case of plants, 13 C-labelling can be achieved by growth in 13 CO 2(g) atmosphere while global 15 N labelling needs 15 N- containing salts in the watering/nutrient solution. Here, we present a method for the preparation of 13 C and 15 N labelled plants by the use of closed growth chambers and hydroponic nutrient supply. The method is exemplified with durum wheat. Results In total, 330 g of globally 13 C- and 295 g of 15 N labelled T. durum wheat was produced during 87 cultivation days. For this, a total of 3.88 mol of 13 CO 2(g) and 58 mmol of 15 N were consumed. The degree of enrichment was determined by LC-HRMS and ranged between 96-98 atom% for 13 C and 95-99 atom% for 15 N, respectively. Additionally, the isotopically labelled plant extracts were successfully used for metabolome-wide internal standardisation of native T.durum plants. Application of an isotope-assisted LC-HRMS workflow enabled the detection of 652 truly wheat-derived metabolites out of which 143 contain N. Conclusion A reproducible cultivation which makes use of climate chambers and hydroponics was successfully adapted to produce highly enriched, uniformly 13 C- and 15 N-labelled wheat. The obtained plant material is suitable to be used in all kinds of isotope-assisted research. The described technical equipment and protocol can easily be applied to other plants to produce 13 C-enriched biological samples when the necessary specific adaptations e.g. temperature and light regime, as well as nutrient supply are considered. Additionally, the 15 N-labelling method can also be carried out under regular glasshouse conditions without the need for customised atmosphere.

scholarly journals Glabralysins, Potential New β-Pore-Forming Toxin Family Members from the Schistosomiasis Vector Snail Biomphalaria glabrata

Genes ◽  
2020 ◽  
Vol 11 (1) ◽  
pp. 65
Author(s):  
Damien Lassalle ◽  
Guillaume Tetreau ◽  
Silvain Pinaud ◽  
Richard Galinier ◽  
Neil Crickmore ◽  
...  
Keyword(s):  
Structure Prediction ◽  
Tertiary Structure ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Biomphalaria Glabrata ◽  
Amino Acid Identity ◽  
New Family ◽  
New Genes ◽  
High Degree ◽  
Trematode Parasites

Biomphalaria glabrata is a freshwater Planorbidae snail. In its environment, this mollusk faces numerous microorganisms or pathogens, and has developed sophisticated innate immune mechanisms to survive. The mechanisms of recognition are quite well understood in Biomphalaria glabrata, but immune effectors have been seldom described. In this study, we analyzed a new family of potential immune effectors and characterized five new genes that were named Glabralysins. The five Glabralysin genes showed different genomic structures and the high degree of amino acid identity between the Glabralysins, and the presence of the conserved ETX/MTX2 domain, support the hypothesis that they are pore-forming toxins. In addition, tertiary structure prediction confirms that they are structurally related to a subset of Cry toxins from Bacillus thuringiensis, including Cry23, Cry45, and Cry51. Finally, we investigated their gene expression profiles in snail tissues and demonstrated a mosaic transcription. We highlight the specificity in Glabralysin expression following immune stimulation with bacteria, yeast or trematode parasites. Interestingly, one Glabralysin was found to be expressed in immune-specialized hemocytes, and two others were induced following parasite exposure.

Plasticity, Hydration and Accessibility in Ribonuclease A. The Structure of a New Crystal Form and its Low-Humidity Variant

1998 ◽  
Vol 54 (6) ◽  
pp. 1343-1352 ◽  
Author(s):  
C. Sadasivan ◽  
H. G. Nagendra ◽  
M. Vijayan
Keyword(s):  
Surface Area ◽  
Tertiary Structure ◽  
Detailed Comparison ◽  
Protein Molecule ◽  
Accessible Surface Area ◽  
Crystal Form ◽  
Ribonuclease A ◽  
Probe Radius ◽  
Low Humidity ◽  
Accessible Surface

The structures of a new crystal form of ribonuclease A and its low-humidity variant, each containing two crystallographically independent molecules, have been determined and refined. A detailed comparison of these structures with those of the other known crystal forms of the enzyme, which have different packing arrangements and solvent composition, leads to a meaningful delineation of the rigid and flexible regions of the protein molecule and the nature of its plasticity. Many of the water molecules which are common to all the structures are involved in bridging different regions of the protein molecule, thus emphasizing the role of water in stabilizing the tertiary structure. The analysis of the structures shows that for a given N or O atom, the level of hydration increases with accessible surface area, but levels off at an area of about 10 Å2. Generally, the hydration level tends to drop when the area increases beyond about 20 Å2. This drop correlates with an increase in the displacement parameter. The analysis also suggests that the van der Waals radii and probe radius normally used in accessible surface area calculations are not appropriate for dealing with all situations.

Cloning and Prokaryotic Expression and Structure Prediction of Prx II the Differentially Expressed Gene under AFB1 Stress

2011 ◽  
Vol 236-238 ◽  
pp. 1078-1082
Author(s):  
Zhen Hong Zhuang ◽  
Feng Zhang ◽  
Yan Yun Li ◽  
Jun Yuan ◽  
Yan Ling Yang ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Prokaryotic Expression ◽  
Structure Prediction ◽  
Tertiary Structure ◽  
Differentially Expressed Gene ◽  
Protein Molecule ◽  
Helix 69 ◽  
Prokaryotic Expression Vector ◽  
Mice Liver ◽  
Α Helix

The gene, prx II, in mice liver was found to be up-regulated under the stress of AFB1 in our previous study. In this study, the total RNA from the mice liver was extracted, and followed by the synthesis of cDNA with the RNA as template by the method of reverse transcription. Then, prx II gene fragment was amplified by PCR with the cDNA as template. After double-digestion and ligation reaction, the prokaryotic expression vector pET28a (+)-prx II was successfully constructed and was transformed into E. coli BL21 (DE3). The protein- Prx II was successfully expressed when induced byIPTG, and finally, Prx II was purified by Ni2+-NTA affinity chromatography. The molecule structure of Prx II, including its signal peptide, hydrophobicity, and its secondary and tertiary structure, was predicted by bioinformatics analysis. The results showed that no signal peptide was predicted in the molecule of Prx II; Five hydrohpobic domains were predicted in the protein molecule, and the average predictive value for its hydrophobicity was -0.151; There were 35% α-helix (69 residues) and 21% β-pleated sheet (42 residues) in the molecule; The tertiary structure of the protein was constituted by seven α-helices and seven β-pleated sheets.

The stereochemical code and the logic of a protein molecule

1969 ◽  
Vol 2 (1) ◽  
pp. 65-92 ◽  
Author(s):  
A. M. Liquori
Keyword(s):  
Tertiary Structure ◽  
Polypeptide Chain ◽  
Water Solution ◽  
Storage System ◽  
Three Dimensional ◽  
Protein Molecule ◽  
Information Storage ◽  
Chemical Information ◽  
Amino Acid Residues ◽  
Surrounding Water

Like DNA and the various forms of RNA, a protein molecule is an information storage system. It contains in fact one or more polypeptide chains which may be regarded as linear sequences of twenty different types of monomer units. It is now clearly established that the chemical information corresponding to a given sequence of a polypeptide chain, containing n amino acid residues, is stored in a segment of one of the two strands of DNA containing 3n nucleotides. The transfer of such information from a gene to a polypeptide chain takes place according to the well-known process involving a transcription and a chemical translation step. This last step leads to a polymer which, in appropriate conditions, takes a three-dimensional conformation or tertiary structure which should correspond to a free-energy minimum of the molecule and its surrounding water solution.

Differentiation of Electrophoretically Similar Hemoglobins—Such as S, D, G, and P; or A2, C, E, and O—by Electrophoresis of the Globin Chains

1974 ◽  
Vol 20 (9) ◽  
pp. 1111-1115 ◽  
Author(s):  
Rose G Schneider
Keyword(s):  
Cellulose Acetate ◽  
Tertiary Structure ◽  
Globin Chain ◽  
Globin Chains ◽  
Molecular Charge ◽  
High Degree ◽  
Secondary And Tertiary Structure ◽  
Combined Data

Abstract Electrophoresis of the globin chains of hemoglobin on cellulose acetate in both acidic and alkaline buffers (pH about 6 and 9) containing urea and 2-mercaptoethanol is a simple, rapid means of characterizing hemoglobins. Erythrocyte hemolysate is electrophoresed in the presence of a large amount of mercaptoethanol, which liberates heme from globin, and keeps it in solution during its rapid electrophoretic removal. Each globin chain migrates at a characteristic rate, which varies with the pH and composition of the buffer. The combined data permit differentiation, with a high degree of specificity, of some similarly charged hemoglobins. They may also be useful in assessing the effect of secondary and tertiary structure on molecular charge.

Preparation of uniformly labelled 13C- and 15N-plants using customised growth chambers

2020 ◽  
Author(s):  
Asja Ceranic ◽  
Maria Doppler ◽  
Christoph Büschl ◽  
Alexandra Parich ◽  
Kangkang Xu ◽  
...  
Keyword(s):  
Plant Material ◽  
Life Science ◽  
Plant Stress ◽  
Science Research ◽  
Nutrient Supply ◽  
Light Regime ◽  
Technical Equipment ◽  
Growth Chambers ◽  
High Degree ◽  
Labelling Method

Abstract Background Stable isotopically labelled organisms found wide application in life science research including plant physiology, plant stress and defense as well as metabolism related sciences. Therefore, the reproducible production of plant material enriched with stable isotopes such as 13 C and 15 N is of considerable interest. A high degree of enrichment (>96 atom%) with a uniformly distributed isotope (global labelling) is accomplished by a continuous substrate supply during plant growth/cultivation. In the case of plants, 13 C-labelling can be achieved by growth in 13 CO 2(g) atmosphere while global 15 N labelling needs 15 N- containing salts in the watering/nutrient solution. Here, we present a method for the preparation of 13 C and 15 N labelled plants by the use of closed growth chambers and hydroponic nutrient supply. The method is exemplified with durum wheat.Results In total, 330 g of globally 13 C- and 295 g of 15 N labelled T. durum wheat was produced during 87 cultivation days. For this, a total of 3.88 mol of 13 CO 2(g) and 58 mmol of 15 N were consumed. The degree of enrichment was determined by LC-HRMS and ranged between 96-98 atom% for 13 C and 95-99 atom% for 15 N, respectively. Additionally, the isotopically labelled plant extracts were successfully used for metabolome-wide internal standardisation of native T.durum plants. Application of an isotope-assisted LC-HRMS workflow enabled the detection of 652 truly wheat-derived metabolites out of which 143 contain N.Conclusion A reproducible cultivation which makes use of climate chambers and hydroponics was successfully adapted to produce highly enriched, uniformly 13 C- and 15 N-labelled wheat. The obtained plant material is suitable to be used in all kinds of isotope-assisted research. The described technical equipment and protocol can easily be applied to other plants to produce 13 C-enriched biological samples when the necessary specific adaptations e.g. temperature and light regime, as well as nutrient supply are considered. Additionally, the 15 N-labelling method can also be carried out under regular glasshouse conditions without the need for customised atmosphere.

Bottom-Up Mesoscale Model of Microtubule

2007 ◽  
Author(s):  
Marco A. Deriu ◽  
Søren Enemark ◽  
Emiliano Votta ◽  
Franco M. Montevecchi ◽  
Alberto Redaelli ◽  
...  
Keyword(s):  
Hollow Cylinder ◽  
Building Block ◽  
Tertiary Structure ◽  
Mesoscale Model ◽  
Eukaryotic Cells ◽  
Outer Diameter ◽  
Structural Elements ◽  
High Degree ◽  
Degree Of Similarity ◽  
Β Tubulin

Microtubules (MTs) are fundamental structural elements in the cytoskeleton of all eukaryotic cells. The MTs are hollow cylinder-shaped biopolymers with inner and outer diameter of about 18 and 30 nm respectively and length ranging from 1 to 10 μm. They are constituted by αβ-tubulins arranged in protofilaments with head-to-tail motif. The protofilaments bind together laterally along the MT’s long axis with a slight shift generating a spiral with a pitch of 2, 3 or 4 monomers’ length [1]. The building-block of the MT, αβ-tubulin, is a hetero-dimer made of two globular monomers, α- and β-tubulin. α- and β-tubulin monomers consists of about 450 residues and shows a high degree of similarity from the primary to the tertiary structure level. However, one important difference is that the α-monomer binds a GTP molecule while the β-monomer binds a GDP molecule [2].

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