scholarly journals Identification of the apparently essential lysine residues in phospholipase C (Bacillus cereus)

1981 ◽  
Vol 193 (3) ◽  
pp. 805-809
Author(s):  
B J Myrnes ◽  
C Little
Keyword(s):  
Amino Acid ◽  
Bacillus Cereus ◽  
Phospholipase C ◽  
Primary Structure ◽  
Lysine Residue ◽  
The Other ◽  
Proline Residue ◽  
Terminal Fragment ◽  
Lysine Residues ◽  
Essential Lysine Residues

Phospholipase C (Bacillus cereus) contains two apparently essential and very reactive lysine residues that may be labelled selectively by pyridoxal 5′-phosphate [Aurebekk & Little (1977) Biochem, J. 161, 159–165]. One of these lysine residues was found in the 25-amino acid N-terminal fragment liberated by CNBr digestion of the pyridoxal-labelled enzyme and identified as lysine-6. Two of the labelled peptides isolated from the chymotryptic digest of pyridoxal-labelled enzyme contained proline, suggesting that the other labelled lysine residue is situated in the same region of the primary structure as the single proline residue of the enzyme.

Die primäre Proteinstruktur von Stämmen des Tabakmosaikvirus

1969 ◽  
Vol 24 (7) ◽  
pp. 870-877 ◽  
Author(s):  
J. Jauregui-Adell ◽  
I. Hindennach ◽  
H. G. Wittmann
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Coat Protein ◽  
Tobacco Mosaic Virus ◽  
Primary Structure ◽  
Amino Acid Sequences ◽  
The Other ◽  
Protein Subunit ◽  
Tryptic Peptides ◽  
Tmv Strains

The sequence of amino acids within the coat protein of the strain Holmes rib grass of tobacco mosaic virus (TMV) has been determined. In this communication the amino acid compositions of the coat protein and of all tryptic peptides are reported. Furthermore the experimental details are given for the elucidation of the amino acid sequences within the first three tryptic peptides, containing 61 amino acids.It has been found that the strain Holmes rib grass differs very extensively in the primary structure from the other TMV strains whose sequences are known. It differs from each of the other strains in more than 50% of the amino acid positions and it contains two amino acids less per protein subunit than the other TMV strains.

scholarly journals The primary structure of histone H2A from the nematode Caenorhabditis elegans

1987 ◽  
Vol 243 (1) ◽  
pp. 297-300 ◽  
Author(s):  
J R Vanfleteren ◽  
S M Van Bun ◽  
J J Van Beeumen
Keyword(s):  
Amino Acid ◽  
Caenorhabditis Elegans ◽  
Primary Structure ◽  
Amino Acid Residues ◽  
Histone H2a ◽  
Nematode Caenorhabditis Elegans ◽  
Calf Thymus ◽  
N Terminus ◽  
Lysine Residues ◽  
Complete Primary Structure

The complete primary structure of histone H2A from the nematode Caenorhabditis elegans was determined. The amino acid chain consists of 126 amino acid residues and has a blocked N-terminus. By comparison with calf thymus histone H2A, the nematode protein shows five deletions, two insertions and 16 substitutions. Most of the changes occur in the N- and C-terminal regions of the molecule, whereas the central part covering the residues 21-120 is quite well conserved. The lysine residues 5, 8 and 10 were found to be partially acetylated.

Comparative study of rye and thymus histones: amino acid analysis and tryptic fingerprinting

1977 ◽  
Vol 55 (7) ◽  
pp. 721-727 ◽  
Author(s):  
Paul Nadeau ◽  
Dominick Pallotta ◽  
Jean-G. Lafontaine
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Secale Cereale ◽  
Primary Structure ◽  
Amino Acid Analysis ◽  
General Pattern ◽  
The Other ◽  
Amino Acid Residues ◽  
Additional Segment ◽  
Similar Amino Acid

Amino acid composition and tryptic fingerprints of rye (Secale cereale) H1, H2B (PHI), and H2A (PHII) histones indicate the presence of major differences between these and the corresponding calf or rabbit fractions. In addition to variations for other amino acids, fraction H1 from rye contains twice as much arginine as the corresponding animal fraction; the plant H2B (PHI) and H2A (PHII) histones show lysine to arginine ratios greater than those of their animal counterparts. The tryptic maps of the same proteins appear to differ between plants and animals by the number and the general pattern of the peptides, as well as by the quantity and distribution of the arginine-containing peptides. Such results suggest the presence of differences in the primary structure of the calf and rye lysine-rich and moderately lysine-rich histones. Furthermore, the possibility is ruled out that each of these plant histones consists of an animal-like protein with an additional segment of 20–30 amino acid residues. On the other hand, the rye and calf arginine-rich fractions H3 and H4 show similar amino acid compositions and tryptic peptides maps.

Active sites of β-lactamases from Bacillus cereus

1980 ◽  
Vol 289 (1036) ◽  
pp. 333-344 ◽  
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Amino Acid ◽  
Bacillus Cereus ◽  
Active Site ◽  
Metal Ion ◽  
Active Sites ◽  
Extracellular Enzyme ◽  
Thiol Group ◽  
The Other ◽  
Site Group

There are two extracellular β-lactamases produced by Bacillus cereus 569. One of these enzymes, β-lactamase I, is inactivated by 6-β-bromopenicillanic acid: the site of reaction is serine-44. This is a conserved amino acid residue in the other β-lactamases whose structures have been determined, and it becomes a good candidate for an active-site group in these enzymes. The inactivation may involve a rearrangement leading to a dihydrothiazine. The other extracellular enzyme produced by B. cereus , β- lactamase II, is exceptional in requiring metal ions for activity. The Zn II and Co II enzymes (the former is more active) have been studied by nuclear magnetic resonance, and by absorption spectroscopy. The groups that bind the metal ion required for activity are three histidine residues and the enzyme’s sole thiol group.

scholarly journals Characterization of a Novel Thrombin-like Enzyme, Globlase from the Venom of Gloydius blomhoffii (Japanese Mamushi)

2019 ◽  
pp. 1-8
Author(s):  
Yumiko Komori ◽  
Yusuke Takashima ◽  
Shoko Ono ◽  
Toshiaki Nikai
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Mass ◽  
Isoelectric Point ◽  
Primary Structure ◽  
Polyacrylamide Gel Electrophoresis ◽  
Hydrolytic Activity ◽  
The Other ◽  
Clotting Factor ◽  
Human Fibrinogen

Aims: To elucidate the coagulation mechanisms of a novel clotting factor isolated from Gloydius blomhoffii venom, its hydrolytic activity on various substrates were examined. Furthermore the primary structure was determined and compared with the other snake venom components. Methodology: A thrombin-like enzyme was isolated from the crude venom of G. blomhoffii by DE52 Cellulose and CM52 Cellulose column chromatography. Enzyme activity was measured by using synthetic substrates (arginine esters, MCA-substrates and 3-(Acyloxy)-4-nitrobenzoic acid). Effect on fibrinogen was detected with bovine and human fibrinogen. Isoelectric point and molecular mass were measured by polyacrylamide gel electrophoresis and MALDI-TOF-MS. Amino acid sequence was decided with a protein sequencer by analyzing enzymatically cleaved peptides. Results: A clotting factor was found to be homologous as indicated by a single band on SDS-PAGE, and the final preparation was named as globlase. Molecular mass of this enzyme was determined to be 13,876.36 Da and the isoelectric point was 8.8. Globlase showed arginine ester hydrolytic activity, and specificity for substrates of thrombin. Proteolytic activity and phospholipase A2 (PLA2) activity were not detected. Complete amino acid sequence analysis indicated that the primary structure of globlase is similar to PLA2. However the aspartic acid which exists in the active site of PLA2 was found to be substituted by glutamine. Conclusion: It was shown in our current investigation that globlase is a novel thrombin-like enzyme isolated from G. blomhoffii venom. It was revealed that this enzyme had structure unlike the serine-protease such as the other thrombin-like enzymes.

scholarly journals Rapid removal of acetimidoyl groups from proteins and peptides. Applications to primary structure determination

1981 ◽  
Vol 199 (2) ◽  
pp. 335-340 ◽  
Author(s):  
G C Dubois ◽  
E A Robinson ◽  
J K Inman ◽  
R N Perham ◽  
E Appella
Keyword(s):  
Amino Acid ◽  
Primary Structure ◽  
Half Life ◽  
Structural Studies ◽  
Amino Groups ◽  
Mouse Immunoglobulin ◽  
Lysine Residues ◽  
Rapid Removal ◽  
Proteins And Peptides

Methylamine buffers can be used for the rapid quantitative removal of acetimidoyl groups from proteins and peptides modified by treatment with ethyl or methyl acetimidate. The half-life for displacement of acetimidoyl groups from fully amidinated proteins incubated in 3.44 M-methylamine/HCl buffer at pH 11.5 and 25 degrees C was approx. 26 min; this half life is 29 times less than that observed in ammonia/HCl buffer under the same conditions of pH and amine concentration. Incubation of acetimidated proteins with methylamine for 4 h resulted in greater than 95% removal of acetimidoyl groups. No deleterious effects on primary structure were detected by amino acid analysis or by automated Edman degradation. Reversible amidination of lysine residues, in conjunction with tryptic digestion, has been successfully applied to the determination of the amino acid sequence of an acetimidated mouse immunoglobulin heavy chain peptide. The regeneration of amino groups in amidinated proteins and peptides by methylaminolysis makes amidination a valuable alternative to citraconoylation and maleoylation in structural studies.

N-Terminale Sequenz einer Porphobilinogen-Synthase * N-Terminal Sequence of a Porphobilinogen-Synthase

1983 ◽  
Vol 38 (11-12) ◽  
pp. 1059-1061 ◽  
Author(s):  
Birgit Lingner ◽  
Traute Kleinschmidt
Keyword(s):  
Amino Acid ◽  
Amino Acid Composition ◽  
Primary Structure ◽  
Bovine Liver ◽  
The Other ◽  
Terminal Sequence ◽  
Terminal Amino Acid ◽  
Porphobilinogen Synthase ◽  
Terminal Amino ◽  
Tryptic Hydrolysis

The N-terminal sequence of porphobilinogen-synthase (EC 4.2.1.24) from bovine liver up to position 44 is given. After tryptic hydrolysis two N-terminal peptides with identical amino acid composition were isolated in the ratio 1:1. One peptide was blocked whereas the other one started with methionine and showed the same primary structure which had been estimated by automatical degradation of the enzyme. Four of the eight subunits of PBG-S have free methionine as N-terminal amino acid whereas the other half is blocked

scholarly journals The primary structure of aspartate aminotransferase from pig heart muscle. Digestion with a proteinase having specificity for lysine residues

1975 ◽  
Vol 149 (3) ◽  
pp. 497-506 ◽  
Author(s):  
S Doonan ◽  
H J Doonan ◽  
R Hanford ◽  
C A Vernon ◽  
J M Walker ◽  
...  
Keyword(s):  
Primary Structure ◽  
Aspartate Aminotransferase ◽  
The Other ◽  
British Library ◽  
Side Chains ◽  
Amino Acid Residues ◽  
Main Paper ◽  
Lysine Residues ◽  
Arginine Residues ◽  
Complete Primary Structure

Carboxymethylated aspartate aminotransferase was digested with a proteinase claimed to be specific for lysine residues. Complete cleavage occurred at 12 of the 19 lysine residues in the protein, but at the remaining seven residues cleavage was either restricted or absent. In addition, cleavage was observed at three of the 26 arginine residues. These results are discussed with reference to the amino acid residues adjacent to points of complete or restricted cleavage. The complete primary structure of aspartate aminotransferase, based on these and other studies, is given. Evidence for the assignment of some acid and amide side chains has been deposited as Supplementary Publication SUP 50050 (11 pp.) at the British Library (Lending Division), Boston Spa, Wetherby, W. Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1975) 145, 5. The evidence for the assignment of residue 366 was less conclusive than for the other acid and amide side chains and is, therefore, given in the main paper.

scholarly journals Phospholipase C from Bacillus cereus. Evidence for essential lysine residues

1977 ◽  
Vol 161 (1) ◽  
pp. 159-165 ◽  
Author(s):  
B Aurebekk ◽  
C Little
Keyword(s):  
Enzyme Activity ◽  
Phospholipase C ◽  
Specific Activity ◽  
Schiff’S Base ◽  
Absorbance Maximum ◽  
Complete Inactivation ◽  
Lysine Residues ◽  
Essential Lysine Residues ◽  
Base Formation ◽  
Fluorescent Component

1. Phospholipase C was inactivated by exposure to the three amino-group reagents, ethyl acetamidate, 2,4,6-trinitrobenzensulphonic acid and pyridoxal 5′-phosphate plus reduction. 2. Inactivation by pyridoxal 5′-phosphate showed the characteristics of Schiff's base formation with the enzyme. The pyridoxal 5′-phosphate-treated enzyme after reduction had an absorbance maximum at 325 mm and 6-N-pyridoxyl-lysine was the only fluorescent component after acid hydrolysis. 3. For complete inactivation, 2 mol of pyridoxal 5′-phosphate or 7 mol of 2,4,6-trinitrophenyl were incorporated/mol of enzyme. 4. The two apparently essential lysine residues were much more reactive to pyridoxal 5′-phosphate than the other 19 lysine residues in the enzyme. 5. Binding of phospholipase C to a substrate-based affinity gel caused marked protection against inactivation by pyridoxal 5′-phosphate. For complete inactivation of the gel-bound enzyme, 5 mol of pyridoxal 5′-phosphate were incorporated/mol of enzyme and there was no evidence of two especially reactive lysine residues. 6. On application of pyridoxal 5′-phosphate-treated enzyme (remaining activity 30% of original) to a column of the affinity gel, some material bound and some did not. The latter contained very little enzyme activity and was heavily incorporated with reagent (9.06 mol/mol of enzyme). The former had a specific activity of 34% of that of the control and contained 1.29 mol of reagent/mol of enzyme. 7. Thus phospholipase C appears to contain two lysine residues that are essential for enzyme activity, but probably not for substrate binding.

scholarly journals The primary structure of troponin T and the interaction with tropomyosin

1975 ◽  
Vol 151 (1) ◽  
pp. 85-97 ◽  
Author(s):  
P Jackson ◽  
G W Amphlett ◽  
S V Perry
Keyword(s):  
Skeletal Muscle ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Primary Structure ◽  
Troponin T ◽  
Troponin C ◽  
The Other ◽  
Tryptic Digestion ◽  
Partial Digestion ◽  
Common Sequence

1. Eight peptides were separated from the CNBr digest of troponin T from rabbit white skeletal muscle and characterized. 2. By study of the amino acid sequence of the methionine-containing peptides isolated after chymotryptic and tryptic digestion and of the N- and C-terminals of the CNBr peptides, six of the latter were shown to be arranged in the sequence CNB1-CNB2-CNB5-CNB6-CNB8-CNB7. The other two peptides, CNB1′ and CNB3, have been shown to be partial digestion products. 3. The CNBr peptides CNB1′ and CNB2 contained a common sequence and were the only peptides in CNBr digests of troponin T that formed a complex with tropomyosin as judged by viscometric and electrophoretic studies. 4. It is concluded that tropomyosin interacts with the N-terminal half of the troponin T molecule approximately in the region lying between residues 70 and 160. 5. Electrophoretic evidence indicates that tropomyosin and troponin C interact with troponin T. 6. None of the major CNBr peptides of troponin T isolated formed a complex with troponin C on electrophoresis at pH 8.6.

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