Development of novel enantioselective HPLC methods for the determination of the optical purity of Nα-Fmoc/Boc amino acid derivatives (Nα-PADs) of natural and unnatural amino acids: an analytical strategy for the key starting materials of therapeutic peptides

2018 ◽  
Vol 10 (21) ◽  
pp. 2481-2493 ◽  
Author(s):  
Yagnakirankumar Komaravolu ◽  
Venugopala Rao Dama ◽  
Thirumala Chary Maringanti
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Unnatural Amino Acids ◽  
Optical Purity ◽  
Amino Acid Derivatives ◽  
Chiral Hplc ◽  
Analytical Strategy ◽  
Therapeutic Peptides

Chiral HPLC methods for natural and unnatural Nα-Fmoc/Boc amino acid derivatives used in therapeutic peptides.

Fluorescence Reagents for Labelling of Biomolecules. Part II. Reactions of 9-Isothiocyanatoacridine with Amino Acids

1994 ◽  
Vol 59 (1) ◽  
pp. 213-221 ◽  
Author(s):  
Dušan Podhradský ◽  
Peter Oravec ◽  
Marián Antalík ◽  
Pavol Kristian
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Structure Determination ◽  
Phenyl Isothiocyanate ◽  
Amino Acid Derivatives ◽  
Analogous Reaction ◽  
Kinetic Measurements ◽  
Fluorescence Labelling

N-(9-Acridinylthiocarbamoyl)amino acids (ATC-AA) II - VII were synthesized by reaction of amino acids with 9-isothiocyanatoacridine I, a new fluorescence labelling agent. The amino acid derivatives II - VII show high relative fluorescence, which is suitable for the determination of nanomolar amounts of ATC-AA. The kinetic measurements show that reaction of I with amino acids is 6 to 22 times faster than analogous reaction of phenyl isothiocyanate. The possibility of using 9-isothiocyanatoacridine for structure determination of proteins is discussed.

A new double-labelling procedure for determination of amino acid composition: Application to bacteriorhodopsin

1978 ◽  
Vol 56 (6) ◽  
pp. 517-520 ◽  
Author(s):  
H. Kaplan ◽  
D. C. H. Cheng ◽  
G. Oda ◽  
M. Kates
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Mixture ◽  
Acid Mixture ◽  
Amino Acid Derivatives ◽  
Double Labelling ◽  
Tryptophan Residues ◽  
Labelling Procedure ◽  
Derivatives Of

A new double-labelling procedure for amino acid analysis which requires only routine chromatographic equipment is described. When 1-fluoro-2,4-dinitro[3H]benzene is reacted with a mixture of 14C-labelled amino acids followed by reaction with the same 14C-labelled amino acid mixture diluted with an unlabelled sample of amino acids, the 3H: 14C ratio in the resulting 2,4-dinitrophenyl (DNP) amino acid derivatives of the diluted sample will be increased in proportion to the quantity of unlabelled amino acid in the diluted sample. This procedure gave reliable results when applied to the known proteins insulin and lysozyme. The procedure is most advantageous when applied to amino acids which are unstable during acid hydrolysis or present in low molar fractions.When applied to the analysis of the bacteriorhodopsin in Halobacterium cutirubrum, this procedure showed the presence of one histidine residue and four tryptophan residues per mole protein but no cystine or cysteine; in general, the analyses obtained were consistent with those originally reported by Oesterhelt, D. and Stoeckenius, W. (1971) (Nature (London) New Biol. 233, 149–152) for bacteriorhodopsin of H. halobium.

scholarly journals Transferability of N-terminal mutations of pyrrolysyl-tRNA synthetase in one species to that in another species on unnatural amino acid incorporation efficiency

Amino Acids ◽  
2020 ◽  
Author(s):  
Thomas L. Williams ◽  
Debra J. Iskandar ◽  
Alexander R. Nödling ◽  
Yurong Tan ◽  
Louis Y. P. Luk ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Genetic Code ◽  
Unnatural Amino Acids ◽  
Trna Synthetase ◽  
Unnatural Amino Acid ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Genetic Code Expansion ◽  
Incorporation Efficiency

AbstractGenetic code expansion is a powerful technique for site-specific incorporation of an unnatural amino acid into a protein of interest. This technique relies on an orthogonal aminoacyl-tRNA synthetase/tRNA pair and has enabled incorporation of over 100 different unnatural amino acids into ribosomally synthesized proteins in cells. Pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNA from Methanosarcina species are arguably the most widely used orthogonal pair. Here, we investigated whether beneficial effect in unnatural amino acid incorporation caused by N-terminal mutations in PylRS of one species is transferable to PylRS of another species. It was shown that conserved mutations on the N-terminal domain of MmPylRS improved the unnatural amino acid incorporation efficiency up to five folds. As MbPylRS shares high sequence identity to MmPylRS, and the two homologs are often used interchangeably, we examined incorporation of five unnatural amino acids by four MbPylRS variants at two temperatures. Our results indicate that the beneficial N-terminal mutations in MmPylRS did not improve unnatural amino acid incorporation efficiency by MbPylRS. Knowledge from this work contributes to our understanding of PylRS homologs which are needed to improve the technique of genetic code expansion in the future.

scholarly journals Determination of amino acids in human biological fluids by high-performance liquid chromatography: critical review

Amino Acids ◽  
2021 ◽  
Author(s):  
Grażyna Gałęzowska ◽  
Joanna Ratajczyk ◽  
Lidia Wolska
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
High Performance ◽  
Hplc Analysis ◽  
Limit Of Detection ◽  
Biological Fluids ◽  
Analytical Techniques ◽  
Epidemiological Studies ◽  
Preparation Conditions

AbstractThe quantitation and qualification of amino acids are most commonly used in clinical and epidemiological studies, and provide an excellent way of monitoring compounds in human fluids which have not been monitored previously, to prevent some diseases. Because of this, it is not surprising that scientific interest in evaluating these compounds has resurfaced in recent years and has precipitated the development of a multitude of new analytical techniques. This review considers recent developments in HPLC analytics on the basis of publications from the last few years. It helps to update and systematize knowledge in this area. Particular attention is paid to the progress of analytical methods, pointing out the advantages and drawbacks of the various techniques used for the preparation, separation and determination of amino acids. Depending on the type of sample, the preparation conditions for HPLC analysis change. For this reason, the review has focused on three types of samples, namely urine, blood and cerebrospinal fluid. Despite time-consuming sample preparation before HPLC analysis, an additional derivatization technique should be used, depending on the detection technique used. There are proposals for columns that are specially modified for amino acid separation without derivatization, but the limit of detection of the substance is less beneficial. In view of the fact that amino acid analyses have been performed for years and new solutions may generate increased costs, it may turn out that older proposals are much more advantageous.

scholarly journals Determination of Methylated Basic Amino Acids with the Amino Acid Analyzer

1973 ◽  
Vol 248 (7) ◽  
pp. 2387-2391 ◽  
Author(s):  
Gladys E. Deibler ◽  
Russell E. Martenson
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Basic Amino Acids ◽  
Amino Acid Analyzer

scholarly journals Determination of free amino acids in plants by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS)

2015 ◽  
Vol 7 (18) ◽  
pp. 7574-7581 ◽  
Author(s):  
Magdalena M. Dziągwa-Becker ◽  
Jose M. Marin Ramos ◽  
Jakub K. Topolski ◽  
Wiesław A. Oleszek
Keyword(s):  
Mass Spectrometry ◽  
Amino Acids ◽  
Liquid Chromatography ◽  
Amino Acid ◽  
Tandem Mass Spectrometry ◽  
Free Amino ◽  
Tandem Mass ◽  
Amino Acid Determination ◽  
Acid Determination

Free amino acid determination in plants by LC-MS/MS.

The Use of Chiral Recognition in a Bioorganic System for Determination of Optical Purity of Labelled Amino Acids

2010 ◽  
Vol 91 (5) ◽  
pp. 491-491
Author(s):  
G. Bergson ◽  
C. Halldin ◽  
H. Lundqvist ◽  
B. Långström ◽  
M. Malmqvist
Keyword(s):  
Amino Acids ◽  
Chiral Recognition ◽  
Optical Purity

scholarly journals Engineered unnatural ubiquitin for optimal detection of deubiquitinating enzymes

2020 ◽  
Author(s):  
Wioletta Rut ◽  
Mikołaj Żmudziński ◽  
Scott J. Snipas ◽  
Miklos Bekes ◽  
Tony T. Huang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Neurodegenerative Disorders ◽  
Unnatural Amino Acids ◽  
Biochemical Analysis ◽  
Deubiquitinating Enzymes ◽  
Fluorogenic Substrates ◽  
Protein Conjugates ◽  
Highly Active ◽  
Selective Chemical

AbstractDeubiquitinating enzymes (DUBs) are responsible for removing ubiquitin (Ub) from its protein conjugates. DUBs have been implicated as attractive therapeutic targets in the treatment of viral diseases, neurodegenerative disorders and cancer. The lack of selective chemical tools for the exploration of these enzymes significantly impairs the determination of their roles in both normal and pathological states. Commercially available fluorogenic substrates are based on the C-terminal Ub motif or contain Ub coupled to a fluorophore (Z-LRGG-AMC, Ub-AMC); therefore, these substrates suffer from lack of selectivity. By using a hybrid combinatorial substrate library (HyCoSuL) and a defined P2 library containing a wide variety of nonproteinogenic amino acids, we established a full substrate specificity profile for two DUBs—MERS PLpro and human UCH-L3. Based on these results, we designed and synthesized Ub-based substrates and activity-based probes (ABPs) containing selected unnatural amino acids located in the C-terminal Ub motif. Biochemical analysis and cell-based experiments confirmed the activity and selectivity of engineered Ub-based substrates and probes. Using this approach, we propose that for any protease that recognizes Ub and Ub-like substrates, a highly active and selective unnatural substrate or probe can be engineered.

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