Attempting to rewrite History: challenges with the analysis of histidine-phosphorylated peptides

2013 ◽  
Vol 41 (4) ◽  
pp. 1089-1095 ◽  
Author(s):  
Maria-Belen Gonzalez-Sanchez ◽  
Francesco Lanucara ◽  
Matthew Helm ◽  
Claire E. Eyers
Keyword(s):  
Amino Acids ◽  
Protein Function ◽  
Biologically Relevant ◽  
Phosphorylated Peptides ◽  
Higher Eukaryotes ◽  
Histidine Phosphorylation ◽  
Rapid Hydrolysis ◽  
Hydrolysis Of ◽  
Acid Labile

A significant number of proteins in both eukaryotes and prokaryotes are known to be post-translationally modified by the addition of phosphate, serving as a means of rapidly regulating protein function. Phosphorylation of the amino acids serine, threonine and tyrosine are the focus of the vast majority of studies aimed at elucidating the extent and roles of such modification, yet other amino acids, including histidine and aspartate, are also phosphorylated. Although histidine phosphorylation is known to play extensive roles in signalling in eukaryotes, plants and fungi, roles for phosphohistidine are poorly defined in higher eukaryotes. Characterization of histidine phosphorylation aimed at elucidating such information is problematic due to the acid-labile nature of the phosphoramidate bond, essential for many of its biological functions. Although MS-based strategies have proven extremely useful in the analysis of other types of phosphorylated peptides, the chromatographic procedures essential for such approaches promote rapid hydrolysis of phosphohistidine-containing peptides. Phosphate transfer to non-biologically relevant aspartate residues during MS analysis further complicates the scenario.

scholarly journals Avoided motifs: short amino acid strings missing from protein datasets

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Pablo Mier ◽  
Miguel A. Andrade-Navarro
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Protein Function ◽  
Large Protein ◽  
New Approach ◽  
Cellular Context ◽  
Human Proteins ◽  
Context Specific ◽  
Protein Datasets

Abstract According to the amino acid composition of natural proteins, it could be expected that all possible sequences of three or four amino acids will occur at least once in large protein datasets purely by chance. However, in some species or cellular context, specific short amino acid motifs are missing due to unknown reasons. We describe these as Avoided Motifs, short amino acid combinations missing from biological sequences. Here we identify 209 human and 154 bacterial Avoided Motifs of length four amino acids, and discuss their possible functionality according to their presence in other species. Furthermore, we determine two Avoided Motifs of length three amino acids in human proteins specifically located in the cytoplasm, and two more in secreted proteins. Our results support the hypothesis that the characterization of Avoided Motifs in particular contexts can provide us with information about functional motifs, pointing to a new approach in the use of molecular sequences for the discovery of protein function.

Iron(III) complexes of DL-methionine

1972 ◽  
Vol 25 (2) ◽  
pp. 421 ◽  
Author(s):  
EJ Halbert ◽  
MJ Rogerson
Keyword(s):  
Aqueous Solution ◽  
Amino Acids ◽  
Large Excess ◽  
Ph Titration ◽  
Oxidation Reduction ◽  
Infrared Studies ◽  
Rapid Hydrolysis ◽  
Iron Analysis ◽  
Hydrolysis Of ◽  
Made In

Few iron(111) complexes of amino acids have been isolated although attempts have been made to measure their stabilities in solution. Iron(111) in the presence of various amino acids showed no complex formation during pH titration in aqueous solution. Using oxidation-reduction and spectrophotometric techniques Perrin measured the stabilities of 1 : 1 complexes of iron(111) with different amino acids in solutions of low pH. Rapid hydrolysis of iron(111) occurred when the pH was increased even in the presence of a large excess of amino acids. Bielig and Bayer reported the isolation of a bis-complex of iron(111) and methionine prepared in aqueous solution. McAuliffe, Quagliano, and Vallarinoe reported a tris-complex made in ethanol, though their iron analysis was not consistent with the required structure. In both cases magnetic and infrared studies were used to characterize the products. Our attempts to repeat these preparations were unsuccessful. This paper describes the preparation of two 1 : 1 complexes of methionine and iron(111) and an investigation of their properties by analysis of infrared, magnetic, and N.M.R. measurements.

Isolation and partial characterization of a peptidase with trypsin-like activity from bovine adenohypophysis

1989 ◽  
Vol 54 (8) ◽  
pp. 2276-2286
Author(s):  
Tsezengijn Dash ◽  
Tomislav Barth ◽  
Jiřina Slaninová ◽  
Jana Barthová ◽  
Hana P. Mašková ◽  
...  
Keyword(s):  
Amino Acids ◽  
Molecular Weight ◽  
Partial Characterization ◽  
Chromogenic Substrate ◽  
Fluorogenic Substrate ◽  
Basic Amino Acids ◽  
Optimum Ph ◽  
Reproducible Method ◽  
Hydrolysis Of

A reproducible method has been developed for the isolation of the adenohypophyseal enzyme with a trypsin-like activity. The enzyme is able to hydrolyze Nα-benzoyl-L-arginine-p-nitroanilide, a fluorogenic substrate CBzl-Arg-Arg-β-naphthyl amide and some peptides with one or two accumulated basic amino acids in the chain. The optimum pH for hydrolysis of the chromogenic substrate was within the range 6.0-7.0 (Km = 0.66 mmol l-1), in the case of the fluorogenic substrate the range was between 7.0 and 7.5 (Km = 1.2 μmol l-1). The enzyme is activated by cysteine and dithiothreitol and inhibited by SH-poisons. The molecular weight of the enzyme, determined by means of two independent methods, was approximately 25 kDA.

Studies of Australian Ascidians. IV. Mollamide, a Cytotoxic Cyclic Heptapeptide From the Compound Ascidian Didemnum molle

1994 ◽  
Vol 47 (1) ◽  
pp. 61 ◽  
Author(s):  
AR Carroll ◽  
BF Bowden ◽  
JC Coll ◽  
DCR Hockless ◽  
BW Skelton ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cyclic Peptide ◽  
Peptide Hydrolysis ◽  
One Dimensional ◽  
Crystallographic Study ◽  
Relative Stereochemistry ◽  
Cyclic Heptapeptide ◽  
Hydrolysis Of ◽  
Absolute Stereochemistry

A new cytotoxic cyclic heptapeptide, mollamide, has been isolated from the compound ascidian Didemnum molle. The structure of mollamide was initially deduced from one-dimensional and two-dimensional n.m.r . experiments at 300 MHz. The structure was confirmed by an X-ray crystallographic study, which showed the relative stereochemistry of the cyclic peptide. Hydrolysis of mollamide , and characterization of the amino acids by derivatization with Nα -(5-fluoro-2,4-dinitrophenyl)-L- alaninamide (FDAA; Marfey's reagent) gave the absolute stereochemistry; all constituent amino acids were the L-isomers. Cytotoxicity data are reported.

scholarly journals Deacylation of acylamino compounds other than penicillins by the cell-bound penicillin acylase of Escherichia coli

1969 ◽  
Vol 115 (4) ◽  
pp. 741-745 ◽  
Author(s):  
M. Cole
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Small Group ◽  
Penicillin Acylase ◽  
E Coli ◽  
Rapid Hydrolysis ◽  
Hydrolysis Of

1. The action of the penicillin acylase enzyme of Escherichia coli N.C.I.B. 8743 on non-penicillin substrates suggests that the enzyme is an amidohydrolase. 2. The rates of hydrolysis for a small group of penicillins closely parallel those for a corresponding series of N-acylglycines. 3. For a series of E. coli strains, ability to cause rapid hydrolysis of phenylacetylglycine is correlated with ability to hydrolyse benzylpenicillin. 4. Amides and N-acylglycines are hydrolysed to the corresponding acids. The phenylacetyl group is hydrolysed most readily. Benzamide and β-phenylpropionamide are not substrates. In a series of aliphatic acylglycines only valeryl- and hexanoyl-glycine are substrates. 5. Acylated l- but not d-α-amino acids are hydrolysed. d-α-Hydroxyphenylacetamide is a better substrate than the l compound.

scholarly journals Simultaneous Presence of PrtH and PrtH2 Proteinases inLactobacillus helveticusStrains Improves Breakdown of the Pure αs1-Casein

2010 ◽  
Vol 77 (1) ◽  
pp. 179-186 ◽  
Author(s):  
L. Sadat-Mekmene ◽  
J. Jardin ◽  
C. Corre ◽  
D. Mollé ◽  
R. Richoux ◽  
...  
Keyword(s):  
Amino Acids ◽  
Cell Envelope ◽  
Sodium Dodecyl ◽  
Lactobacillus Helveticus ◽  
Whole Cells ◽  
Proteolytic Activities ◽  
Rapid Hydrolysis ◽  
Milk Casein ◽  
Kinetics Of ◽  
Hydrolysis Of

ABSTRACTLactobacillus helveticuscan possess one or two cell envelope proteinases (CEPs), called PrtH2 and PrtH. The aim of this work was to explore the diversity of 15 strains ofL. helveticus, isolated from various origins, in terms of their proteolytic activities and specificities on pure caseins or on milk casein micelles. CEP activity differed 14-fold when the strains were assayed on a synthetic substrate, but no significant differences were detected between strains possessing one or two CEPs. No correlation was observed between the proteolytic activities of the strains and their rates of acidification in milk. The kinetics of hydrolysis of purified αs1- and β-casein byL. helveticuswhole cells was monitored using Tris-Tricine sodium dodecyl sulfate (SDS) electrophoresis, and for four strains, the peptides released were identified using mass spectrometry. While rapid hydrolysis of pure β-casein was observed for all strains, the hydrolysis kinetics of αs1-casein was the only criterion capable of distinguishing between the strains based on the number of CEPs. Fifty-four to 74 peptides were identified for each strain. When only PrtH2 was present, 22 to 30% of the peptides originated from αs1-casein. The percentage increased to 41 to 49% for strains in which both CEPs were expressed. The peptide size ranged from 6 to 33 amino acids, revealing a broad range of cleavage specificities, involving all classes of amino acids (Leu, Val, Ala, Ile, Glu, Gln, Lys, Arg, Met, and Pro). Regions resistant to proteolysis were identified in both caseins. When strains were grown in milk, a drastic reduction in the number of peptides was observed, reflecting changes in accessibility and/or peptide assimilation during growth.

Improved purification and further characterization of Clostridium perfringens type A enterotoxin

1973 ◽  
Vol 19 (11) ◽  
pp. 1379-1382 ◽  
Author(s):  
A. H. W. Hauschild ◽  
R. Hilsheimer ◽  
W. G. Martin
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Clostridium Perfringens ◽  
Amino Acid Analysis ◽  
Sedimentation Coefficient ◽  
Type A ◽  
Sedimentation Equilibrium ◽  
Molecular Weights ◽  
Hydrolysis Of

The procedure for the purification of Clostridium perfringens type A enterotoxin was simplified, and the purity of the toxin was improved. Hydrolysis of the toxin by the p-toluenesulfonic acid procedure yielded 18 common amino acids. Among these, aspartic acid, serine, leucine, and glutamic acid were the predominant components. The sedimentation coefficient (s°20, w) was 2.8 Svedberg units. The molecular weights determined by the Archibald technique, sedimentation equilibrium, and amino acid analysis were 40 000, 36 000, and 33 000, respectively.

scholarly journals Isolation and characterization of sheep pepsin

1977 ◽  
Vol 161 (2) ◽  
pp. 389-398 ◽  
Author(s):  
P F Fox ◽  
J R Whitaker
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Composition ◽  
Ethyl Ester ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Isolation And Characterization ◽  
Sepharose 4B ◽  
Hydrolysis Of

Sheep pepsin was isolated (approx. 120-fold purification) from aqueous abomasal homogenates by (1) pH fractionation, (2) chromatography on Sepharose 4B-poly-L-lysine columns and (3) gel filtration on Sephadex G-100. The enzyme had mol.wt. approx. 34000, N-terminal valine and C-terminal alanine. The amino acid composition of sheep pepsin was generally similar to that of pig and ox pepsins, with a very low content of basic residues and a high content of acidic and hydroxy-amino acids. The pH optimum for NN-dimethyl-casein and NN-dimethyl-haemoglobin as substrates was approx. 1.8. The Km and kcat. for NN-dimethyl-haemoglobin were 46micronM and 1100min-1 respectively, and for NN-dimethyl-casein the corresponding parameters were 50micronM and 420min-1. These values were generally similar to those for pig and ox pepsins. At the pH optimum of 4.6, the sheep pepsin was about 50% as active on benzyloxycarbonyl-L-histidyl-L-phenyl-alanyl-L-tryptophan ethyl ester as was pig pepsin. The pH optimum for the hydrolysis of N-acetyl-L-phenylalanyl-L-di-iodotyrosine by sheep, ox and pig pepsins was approx. 1.85.

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